vvEPA
             United States
             Environmental Protection
             Agency
            Industrial Environmental Research EPA 600 2 78-207
            Laboratory         September 1978
            Cincinnati OH 45268
             Research and Development
Reuse of
Fermentation
Brines in the
Cucumber Pickling
Industry
\
     600/2
\  78-207
y/A

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                RESEARCH REPORTING SERIES

Research reports of the Office of Research and Development, U S  Environmental
Protection Agency, have been grouped into nine series These nine broad cate-
gories were established to facilitate further development and application of en-
vironmental technology Elimination of traditional grouping was consciously
planned to foster technology transfer and a maximum interface in related fields
The nine series are

      1   Environmental Health  Effects Research
      2   Environmental Protection Technology
      3   Ecological Research
      4   Environmental Monitoring
      5   Socioeconomic Environmental Studies
      6   Scientific and Technical Assessment Reports (STAR)
      7   Interagenoy Energy-Environment Research and Development
      8   "Special" Reports
      9   Miscellaneous Reports

This report has been assigned  to the  ENVIRONMENTAL PROTECTION TECH-
NOLOGY series This series describes researcn performed to develop and dem-
onstrate instrumentation, equipment, and methodology to repair  or prevent en-
vironmental degradation from point and non-point sources of pollution This work
provides the new or improved technology required for the control and treatment
of pollution sources to meet environmental quality standards.
 This document is available to the public through the National Technical Informa-
 tion Service, Springfield, Virginia  22161

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                                                 EPA-600/2-78-207
                                                 September 1978
           REUSE OF FERMENTATION BRINES IN THE
               CUCUMBER PICKLING INDUSTRY
                            by

                R. F. McFeeters and W. Coon
       Department of Food Science and Human Nutrition
  Michigan State University, East Lansing, Michigan  48824
                           and
             M. P. Palnitkar and M. Velting
  Vlasic Foods, Inc., West Bloomfield, Michigan  48033
                           and
                      N. Fehringer
Detroit District Office, U.S. Food and Drug Administration
                Detroit, Michigan  48826
                   Grant No. S-803825
                      Project Officer

                    Harold W. Thompson
               Food and Wood Products Branch
       Industrial Environmental Research Laboratory
                  Corvallis Field Station
                 Corvallis, Oregon  97330
       INDUSTRIAL ENVIRONMENTAL RESEARCH LABORATORY
            OFFICE OF RESEARCH AND DEVELOPMENT
           U.S.  ENVIRONMENTAL PROTECTION AGENCY
                  CINCINNATI, OHIO  45268

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                           DISCLAIMER
     This report has been reviewed by the Industrial Environ-
mental Research Laboratory-Cincinnati Food and Wood Products
Branch, U.S. Environmental Protection Agency, and approved for
publication.  Approval does not signify that the contents
necessarily reflect the views and policies of the U.S. Environ-
mental Protection Agency, nor does mention of trade names or
commercial products constitute endorsement or recommendation
for use.
                                ii

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                                  FOREWORD
     When energy and material resources are extracted, processed, converted,
and used, the related pollutional impacts on our environment and even on our
health often require that new and increasingly more efficient pollution
control methods be used.  The Industrial Environmental Research Laboratory-
Cincinnati (lERL-Ci) assists in developing and demonstrating new and improved
methodologies that will meet these needs both efficiently and economically.

     This report shows the results of a commercial evaluation of the reuse
of "spent brine" in the cucumber pickling industry.  It appears that the
cucumber pickling industry can economically recover and reuse most of the
fermentation brines generated in tank yards.  This can lead to important
reductions in waste generation in this industry without significant changes
in the quality of the products produced.  As a result, this report should
be of interest to the processors in this industry and environmental regula-
tory agencies.  For further information on this project contact the Food and
Wood Products Branch, Industrial Pollution Control Division, Industrial
Environmental Research Laboratory-Cincinnati.
                                      David G. Stephan
                                         Director
                       Industrial Environmental Research Laboratory
                                        Cincinnati
                                     iii

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                            ABSTRACT

     The project evaluated on a commercial scale the technolo-
gical and economic feasibility of recycling spent cucumber
fermentation brine.  Two brine treatment procedures, heat treat-
ment and chemical treatment, were used.  The results showed
that brine recycling was practical on a commercial scale.  Either
brine treatment procedure resulted in salt stock which were
equivalent in quality to control cucumbers.

     Studies were conducted to determine the adequacy of the
brine treatment procedures employed.  The data confirmed that a
heat treatment of 175°F for 30 sec. was sufficient to assure
inactivation of pectinases from molds found to be common on
cucumber fruits and flowers.  For effective chemical treatment
a brine temperature of 72°F or higher was required.  In addition,
the pH had to be maintained at 11.0 or higher for at least 36 hr
to assure 99% inactivation of pectinases from the molds which
were investigated.

     An economic evaluation of the recycling procedures showed a
small net savings for the heat treatment procedure and a small
net cost for chemical treatment.  Selection of the process for
a particular plant will depend upon the local conditions.

     This report was submitted in fulfillment of Grant S-803825
by Pickle Packers International, Inc., under the partial spon-
sorship of the U.S. Environmental Protection Agency.  This
report covers a period from May, 1975 to December, 1977; data
collection was completed as of October, 1977.
                                iv

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                            CONTENTS



Foreword	iii



Abstract	iv



Figures	vi



Tables	vii



Conversion Factors and Metric Prefixes 	  xi



Acknowledgment 	  xiii



    1. Introduction  	   1



    2. Conclusions 	   4



    3. Recommendations 	   5



    4. Experimental Design 	   6



    5. Results and Discussion	23



References	68



Appendix	71
                               v

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                            FIGURES

Number                                                     Page

  1  Schematic diagram of the brine recovery unit ....    8

  2  Regenerative heat exchanger system used for the
       brine recovery unit	    9
     General design of the commercial evaluation
       of brine recycling	   12

     Score sheet for taste panel evaluation of
       hamburger dill chips	   15

     Effect of pH on heat inactivation of
       Penicillium ianthinellum pectinase.
       Experiments were done in brine containing
       12% NaCl, 0.6% lactic acid and 0.1%
       Ca++ ion	   64
                               vi

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                            TABLES

Number
  1  Characteristics of 1st cycle spent brine in
       the spring of 1975 before and after heat
       or chemical treatment.  The treated brines
       were used as cover brines for 2nd cycle
       fermentations 	    24

  2  Characteristics of 2nd cycle spent brine in
       the spring of 1976 before and after heat
       or chemical treatment.  The treated brines
       were used as cover brines for 3rd cycle
       fermentations 	    25

  3  Summary data on fermentation tanks, 1975	    28

  4  Summary data on fermentation tanks, 1976	    28

  5  Analysis of spent brine remaining in the
       fermentation tanks in the winter after removal
       of salt stock cucumbers from the 2nd cycle
       (1975) and 3rd cycle  (1976) fermentation
       tanks	    30

  6  Analysis of spent brine remaining in the
       fermentation tanks in the winter after
       removal of salt stock cucumbers from the
       2nd cycle (1975) and 3rd cycle  (1976)
       fermentation tanks.  All data are
       expressed as mg/1 of brine	    31

  7  Quality of 2nd cycle salt stock cucumbers
       fermented in recycled brine, 1975 	    32

  8  Quality of 3rd cycle salt stock cucumbers
       fermented in recycled brine, 1976 	    32

  9  Comparison of 2nd cycle product with the
       control (averages), 3 samples/group,
       9 judges and 7 groups	    34
                               Vll

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Number
  10  Comparison of 3rd cycle product with the
        control (averages), 3 samples/group,
        11 judges and 5 groups	35

  11  Analysis of variance table for texture
        differences of hamburger dill chips
        made from 2nd cycle salt stock
        cucumbers	36

  12  Analysis of variance table for flavor
        differences of hamburger dill chips
        made from 2nd cycle salt stock
        cucumbers	37

  13  Analysis of variance table for texture
        differences of hamburger dill chips
        made from 3rd cycle salt stock
        cucumbers	38

  14  Analysis of variance table for flavor
        differences of hamburger dill chips
        made from 3rd cycle salt stock
        cucumbers	39

  15  Analysis of variance table for overall
        quality differences of hamburger dill
        chips made from 3rd cycle salt stock
        cucumbers	40

  16  Salt stock cucumber analysis.  Distri-
        bution of mineral levels in commercial
        salt stock during three cycles of
        fermentation.  All data are expressed
        as mg/kg of cucumbers	42

  17  Desalted cucumber analysis.  Distribution
        of mineral levels in desalted cucumbers
        during three cycles of fermentation.
        All data are expressed as mg/kg of
        cucumbers	43

  18  Savings from recycling 1,000 gal of
        spent brine	45

  19  Economic evaluation of chemical treatment
        of spent brines	45

  20  Operating costs and savings using heat
        treatment for spent brines 	  46
                               Vlll

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Number
  21  Net present value calculation for comparison
        of chemical treatment with heat treatment
        of brine.  The difference between treatment
        costs per 1,000 gal for chemical compared
        to heat treatment of brine was $16.43 in
        1976.  For 1.7 million gal this is a total
        difference in treatment cost of 1700 x
        $16.55 = $27,931.  With a maintenance cost
        of $500/yr for the unit the net difference
        is $27,431	48

  22  Raw waste load summary	50

  23  EPA effluent guidelines - 1977	50

  24  EPA effluent guidelines - 1983	51

  25  Theoretical salt use and discharge during
        fermentation, storage, untanking and
        desalting of cucumbers.  Results are
        expressed as the Ib of salt required per
        bu (50 Ib) of fresh cucumbers	53

  26  Recoveries  (%)  of added pesticides	56

  27  Lowest levels at which residues were
        quantitated	57

  28  Effect of heat and NaOH treatment on
        carbaryl in pickle brine 	   59

  29  Effect of heat and NaOH treatment on
        parathion in pickle brine  	   60

  30  Effect of heat and NaOH treatment on
        endosulfan in pickle brine 	   61

  31  Effect of heat or NaOH treatment on PCNB
        and PCA in pickle brine	62

  32  Sources and identity of fungi used for
        determination of thermal stability of
        pectinases in brines 	   63

  33  D-values of pectinases with the greatest
        thermal stability in preliminary
        experiments.   D-values were obtained at
        pH 3.3 in 12% NaCl, 0.6% lactic acid and
        0.1% Ca++ ion	63
                               IX

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N limber                                                    Page

  34  Effect of NaCl concentration on the heat
        inactivation of pectinase from
        Penicillium janthinellum at 75°C	   65

  35  Inactivation of pectinase from Penicillium
        janthinellum at high pH in spent brine.
        Temperature 22°C (71.6°F).  Salt
        concentration 8.0%	   65
                                X

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                  CONVERSION FACTORS AND METRIC  PREFIXESa
                             CONVERSION  FACTORS
 To convert from	

Degree Fahrenheit  (°F)

inch (in)

foot (ft)

gallon (gal)

bushel (bu)

3.8 gal/bu

grain (gr)

ounce (oz)

pound (Ib)

gallons per minute
      (gal/min)

ounces per gallons
      (oz/gal)

pounds per ton
      (Ib/ton)

pounds per 1000 pounds
      (lb/1000 Ib)

tons per year
      (ton/yr)

gallons per ton
      (gal/ton)
       to
Degree Celsius  (°C)

metre (m)

metre (m)

metre3 (m3)

metre3 (m3)

0.408 m3/m3

kilogram  (kg)

kilogram  (kg)

kilogram  (kg)

metre3/second
          (m3/s)

kilogram/metre3
          (kg/m3)

kilogram/kilokilogram
          (kg/kkg)

kilogram/kilokilogram
          (kg/kkg)

kilokilogram/year
          (kkg/yr)

metre3/kilokilogram
          (mVkkg)
Multiply by
t°c =0.56  (tQp-32)

2.54 x 10~2

3.048 x 10'1

3.784 x 10"3

3.524 x 10"2

1.0

6.48 x ID'5

3.11 x ID'2

4.536 x 10"1

6.308 x 10'5


8.218


4.643 x 10"1


1.0


9.074 x 10"1


4.17 x 10~3
                                      XI

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  To convert from _

pounds per gallon
       (lb/gal)

pounds per 1000 gallons
       (lb/1000 lb)

cost per pound
         to
cost per gallon
       ($/gal)

cost per 1000 gallon
       ($/1000 gal)

cost per ton
       ($/ton)
  kilogram/metre3
          (kg/m3)

  kilogram/metre3
          (kg/m3)

  cost/kilogram
          ($/kg)

  cost/metre
          ($/m3)

  cost/metre3
          ($/m3)

  cost/kilokilogram
          ($/kkg)
 Multiply by
 1.1984 x 102


 1.1984 x ID'1


 4.536 x 10"1


 2.642 x 102


 2.642 x ID'1


 1.102
                               METRIC PREFIXES
Prefix       Symbol

kilo           k

centi          c
Multiplication factor

        IO3

        io-2
       Example
2 kg = 2 x IO3 grams

2 cm = 2 x IO-2 metre
aStandard for Metric Practice.  ANSI/ASTM Designation:   E  380-76e,   IEEE Std
 268-1976,  American Society for Testing and Materials,  Philadephia,
 Pennsylvania,  February  1976.   37 pp.
                                      xii

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                        ACKNOWLEDGMENTS

     Pickle Packers International, Inc., appreciates the
cooperation of the Department of Food Science and Human Nutri-
tion, Michigan State University; Vlasic Foods, Inc.; and
the PPI Ecology Committee in the planning, administration and
execution of this project.  The Detroit District Office of the
U.S. Food and Drug Administration was very helpful in providing
assistance on pesticide analysis.

     Mr. Herman Blum's assistance in initiation of the project
and his advice and leadership during the project were particu-
larly valuable.  Mr. Michael Velting, Mr. Richard Bowman,
Mr. Scott Smith, Mr. Maury Kittel, and Mr. Charles Santerre
were other members of Vlasic Foods, Inc., who made significant
contributions to the operation of the project.  The assistance
of plant personnel at Imlay City, Michigan is also acknowledged.

     Pectinase inactivation studies were done by Ms. Suparath
Chavana and Mr. Nicholas Palamidis.  Assistance with cucumber
brining, sample collection, and analysis at MSU was provided
by Mr. Michael Fishel, Ms. Debra Patterson, Ms. Luci Chua and
Ms. Patricia Fodell.  Members of the Michigan State University
Cucumber Research Committee provided transportation, repair
services, and advice throughout the project.

     We further acknowledge Mr. Harold Thompson of the
Industrial Environmental Research Laboratory of the Environ-
mental Protection Agency for his guidance and numerous helpful
suggestions in all phases of the project.
                              xiii

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                           SECTION 1

                         INTRODUCTION

     Per capita pickle consumption in the United States has
shown steady increases since 1930 from 1.07 kg to 3.75 kg in
1976.  There are two major classes of cucumber pickles, fresh
pack and process pack.  Fresh pack products are prepared by
processing fresh cucumbers directly into final products.  The
process pack requires initial fermentation of cucumbers, storage
at high salt concentrations, and subsequent desalting prior to
processing into consumer or institutional products.  In 1975
the process pack operation utilized 206,000 metric tons of green
stock or 64% of the green stock purchased by processors (1).
This distribution of product utilization makes the tank yard
a major source of waste in the industry.  Kirk (2) reported that
spent brine accounted for 8% of the total hydraulic load, 44%
of the BOD and 59% of the salt generated in the manufacture
of process pack products.

     There are several sources of wastes from tank yard opera-
tions.  These include spillage, tank overflow during rainy
periods, leakage from wooden tanks, spent fermentation brine
and "process water" generated during desalting operations.
This project was designed to evaluate recycling of spent brines
as a means for control of this source of waste.

     Usual fermentation practices will result in production of
about 40% by volume spent brine in a fermentation tank.  The
cucumber fermentation is normally done at 6.5% NaCl (3).  How-
ever, when the fermentation is completed, additional salt is
added to insure against deterioration of the fruit during tank
yard storage.  As a result, spent brines usually contain 9-15%
NaCl, the BOD will be 10,000-15,000 mg/1 and the pH will be
3.2-3.5.  These characteristics make biological treatment of
waste relatively difficult.  Discharge of brines produced by
tank yard operations may not be possible in some locations
with limits on total dissolved solids.

     In assessing environmental control strategies available
for managing these brines one may wonder if it is feasible to
either reclaim the salt contained therein, or to recycle the
spent solution.

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     Durkee et al. (4) has developed a procedure for salt
recovery from fermentation brines by evaporation of the water
and incineration of the organic matter.  Salt recovered by this
method has been used successfully in cucumber fermentations (5).
However, this technique has not been adopted commercially due
to the high cost of equipment and energy.

     When considering brine recycling, many concerns arise.
One is the potential for loss of product quality if pectinases
were carried over in the recycled brine at levels sufficient
to soften cucumbers during months of fermentation and storage
in the brine.  It is also possible that products of fermenta-
tion could inhibit subsequent fermentations or cause abnormal
fermentations.  There could also be a buildup of toxic com-
pounds.  The destruction of these products might require incin-
eration or other drastic techniques.  Pathogenic organisms will
not, however, be a problem due to the low pH and high salt
content of the brine.

     Geisman and Henne (6, 7) developed a chemical treatment
procedure for reclaiming spent fermentation brines.  Their
procedure requires raising the pH of spent brine to 10 or 11.
This treatment clarified the brine, since a precipitate formed
which occluded suspended cells.  It was also presumed to
inactivate pectinase activity.  After a 48-hr holding period,
the pH was adjusted to 7 and the brine used as a cover brine
for fermentation.  Normal fermentations resulted from use of
this brine.  This showed that incineration was not required
for reuse of salt and provided a practical alternative
treatment.

     Palnitkar and McFeeters  (8) showed that chemically treated
brine could be used for multiple fermentation cycles without
loss of salt stock quality.  They also suggested that heating
of brine could be used as an alternative treatment and
suggested 175°F for 30 sec. as a reasonable heat treatment.

     Cranfield (9) had previously reported on a system for
pasteurization of brine, saturation of the treated brine by
addition of salt and reuse of the treated brine as a saturated
salt solution.

     The primary objectives of this project were to evaluate
the commercial, technological, and economic feasibility of
brine recycling by both heat and chemical treatments  and to
determine the relative merits of these two procedures.

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     The recycling procedures were compared to controls in
which cucumbers were fermented in fresh salt brine.  The control
fermentations were done accord-ing to normal commercial brining
procedures.  The commercial fermentations were carried out at
the Imlay City, Michigan plant of Vlasic Foods, Inc.

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                           SECTION 2

                          CONCLUSIONS

1. Brine recycling is an effective procedure for reduction of
   waste in the manufacture of salt-stock pickles.

2. Both the heat and chemical treatments for spent  brine are
   practical for use in current commercial tank yard operations.

3. Heating spent brine for 30 sec. at 175°F will assure
   at least 99.98% destruction of pectinases from fungi which
   were found to be common on cucumber fruits and flowers.

4. Raising the pH of -72°F spent brine to 11.2 or higher for
   at least 37 hr will decrease Penicillium janthinellum pecti-
   nase activity in spent brines to <1% of its initial activity,
   This should be an adequate treatment for brine recycling.

5. Salt-stock cucumbers produced from fermentations using
   recycled brines are equivalent to control salt stock in
   bloater losses, texture, and flavor.  Consumers  have not
   detected any significant differences in products prepared
   from salt stock fermented in recycled brine.

6. No significant accumulations of metals or pesticides occur
   as a result of brine recycling.

7. The results show that brine can be reused for at least three
   fermentation cycles on a commercial basis.  There are also
   no indications of adverse effects of recycling on salt-stock
   quality or buildup of toxic constituents.  This  suggests
   that brine can continue to be reused beyond three cycles.

8. Under the conditions of this project, heat treatment of
   brine resulted in a small net savings while there was a
   small net cost for chemical treatment.  The relative
   economics of these two recycling procedures may  vary with
   individual circumstances.  However, the use of recycling is
   an economically feasible means for reducing the  waste from
   tank yard operations.

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                           SECTION 3

                        RECOMMENDATIONS

1. Brine recycling should be adopted by the processed cucumber
   pickling industry as a means to reduce wastes.

2. Both heat and chemical treatments of the spent brine are
   effective from a technical viewpoint.  Selection of a
   procedure can be based upon economic considerations and upon
   the relative compatability of the procedures with each
   particular tank yard operation.

3. Care should be taken to insure proper design of a recycling
   system, proper training of operating personnel and proper
   supervision of the procedure, since improper recycling
   techniques hold the potential for significant economic
   losses.

4. Undertake an evaluation of calcium hydroxide as a partial
   or total replacement of sodium hydroxide in the chemical
   treatment procedure.  The use of calcium hydroxide would
   significantly reduce the accident risks associated with
   this procedure.

5. Evaluate the need to remove the precipitate formed during
   chemical treatment.  If precipitate removal is unnecessary,
   it would reduce brine losses and reduce handling of the
   brine.

6. Efforts should be made to reduce tank leakage and overflow,
   since significant brine losses occur from these sources.

7. A major effort is needed to develop salt-stock storage
   technology, which will allow reduction in the salt levels
   maintained in brining tanks, since wastes generated in
   the desalting operation are second only to spent brine as a
   source  of waste in tank yard operations.

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                          SECTION 4

                      EXPERIMENTAL DESIGN

BRINE TREATMENT PROCEDURES

Chemical Treatment of Brine

     Chemical treatment of spent brine was carried out by the
addition of Food Grade NaOH pellets to a tank of spent brine
(8,000-10,000 gal) to raise the pH of the brine to 11 or
greater.

     The amount of NaOH required for each batch of brine was
determined by titrating a 32-oz, spent brine sample from a
well mixed tank with 10% NaOH solution to a pH of 11.5.
Approximately 80% of the calculated NaOH requirement was added
to the tank and the brine was circulated.  A sample of this
partially adjusted brine was titrated to pH 11.5, and the
NaOH requirement calculated.  This amount of NaOH was added and
the tank was circulated with a pump.  A characteristic change
in color of the spent brine to darker yellow, development of an
ammonia-like odor and formation of a precipitate occurred as
the pH was raised.  This two-step NaOH addition procecure
provides insurance against major errors in the amount of NaOH
added and attainment of the proper final pH.

     A precipitate formed when the brine pH was raised.  This
precipitate settled in the tank.  After settling, the clear
brine was pumped into a clean tank.  The precipitate layer was
discarded.  The pH of the NaOH treated brine was adjusted to
pH 4.5-4.7 with 300 grain vinegar.  The amount of vinegar
required was determined by titration of a sample with acetic
acid.  Addition was done by the same procedure as the addition
of NaOH.  After acidification of the brine, it was held until
it was used as a cover brine.

     Great care must be exercised when using sodium hydroxide
for pH adjustments.  Injuries were prevented by use of the
following precautions:

        1. Only college students who had taken
           introductory chemistry handled the sodium
           hydroxide.

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        2. These people were specifically taught
           the problems and dangers associated with
           handling NaOH.

        3. Protective clothing including wrap around
           goggles, acid-base resistant gloves, and
           rain suit was  required for handling NaOH.

        4. A water hose was turned on when NaOH was
           being handled so any spills could be
           rapidly washed.  Eye wash equipment was
           readily available.

        5. NaOH was slowly added by manual addition
           while the brine tank was mixing to minimize
           the possibility of local high concentra-
           tions of NaOH on the surface of the tank.

Heat Treatment of Spent Brine

     Spent brine was heated with a trailer mounted brine
recovery system developed by the APV Company, Inc. of Tonawanda,
New York and Vlasic Foods.  A schematic diagram of the unit
is shown in figure 1.  Figure 2 shows a drawing of the heat
exchanger system.  The unit has a titanium alloy heat exchanger
to resist brine corrosion, a 200 gal stainless steel holding
tank, and a screening system to remove particulate materials
from the brine before it enters the heat exchanger.

     The heat exchanger heats brine from ambient temperature to
68°C (155°F)  by regeneration from the heated brine.  Heating
from 68°C up to 90.5°C (195°F)  is done with hot water heated
by burning propane.  This system allows up to 60% of the
heating to be obtained by cooling the treated brine to a
temperature of 35°C (95°F).

     The entire unit is mounted on a 6 x 16 ft trailer so it
can be moved between tank rows.  In routine operation it was
most efficient to pump brine from nearby rows of tanks to
minimize the number of times the unit had to be moved.  The
unit can be loaded on a truck and transported to other tank
yard locations.  In this way one unit has been used to serve
more than one location.  The unit has required relatively
little maintenance and does not appear to be deteriorating
significantly even though several million gallons of brine have
been treated.

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     Bottled propane gas was used as the fuel source.  The heat
exchanger had a heat capacity of 50 gal/min.  To provide an
extra margin of safety, the brine in this project was treated
to 190°F for 30 sec.  Whenever feasible, the heating of brine
was done on warmer days (>21°C or 70°F)  since warmer brine
required less fuel for treatment.

     An operator continuously monitored the heating operation
to assure proper treatment of the brine and to protect the
equipment in case of malfunction.  A second person was required
on a part-time basis to assist in pumping brine to the unit.

     The flat-bottom design of current wood tanks made it
impractical to remove all the brine from tanks.  Usually about
a 6 inch layer of brine containing dirt, pickle debris, and
other residues remained.  This brine was discarded through the
bung hole in the bottom of the tanks.  It was necessary to
thoroughly clean the tanks prior to filling with treated brine.
Though the tanks cannot be sterilized and the brine will be
open to the air, the contamination of the brine by pumping
into a dirty tank is aesthetically unpleasing and could result
in considerable yeast growth in the brine before use.  The
brine comes from the heat exchanger at a temperature of 90-100°F,
Therefore, it will not sanitize a poorly cleaned tank prior to
cooling.  Tanks were cleaned by manually scrubbing the tanks
with 0.5 oz/gal calcium hypochlorite solution followed by
washing with hydrated lime and rinsing with fresh water.  If
tanks were not used immediately for brine storage, they were
filled with fresh water to prevent the wood from contracting.

     The treated brine was held at its original pH, usually at
pH 3.2-3.5.  Just prior to use, the pH was raised to 4.5 with
NaOH pellets using the procedure described earlier.  Etchells
et al. (10) have recommended a similar pH adjustment for fresh
brine used in controlled fermentation.  In laboratory
experiments this pH adjustment was found to be necessary in
order to minimize bloating problems and to obtain a normal acid
development during fermentations.  The quantities of NaOH
required for this adjustment have been safely handled in
normal commercial operations using the precautions previously
described.  It would be worthwhile to obtain data on the use
of lime, Ca(OH)2, for this adjustment.  Calcium salts have
been added to pickles as firming agents.  Lime would be
considerably less expensive than NaOH and easier to handle.
                               10

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Controls for Fermentations With Recycled Brine

     Commercial practice for cucumber fermentations has
historically been to cover the fresh cucumbers with freshly
prepared brine.  At the conclusion of fermentation and storage,
the brine remaining after salt stock was removed from tanks was
termed "spent brine" and discarded.  To evaluate the effects of
brine recycling it was necessary to compare the salt stock
and brines from fermentations with recycled brines with fer-
mentations in which green stock cucumbers were covered with
freshly prepared salt solutions.

COMMERCIAL EVALUATION OF BRINE RECYCLING

     Fermentation brines were reclaimed and reused through three
complete fermentation cycles per treatment as shown in figure 3.
The project began in 1975 with the segregation of first cycle
spent brine from the general tank yard brines.  After heating
or chemical treatment, it was used for second cycle fermenta-
tions in 1975.  The second cycle brine was saved and treated
in the spring of 1976 for third cycle fermentations.  Control
fermentations in which cucumbers were covered with fresh salt
brine were run in both 1975 and 1976.

     Size 3A-3B (3.8-5.1 cm diam) mixed cucumbers were used for
all tanks.  These sizes of cucumbers are fermented in greatest
volume commercially.  In addition, the large size cucumbers
are most susceptible to bloating.  One objective of the project
was to determine if brine recycling would significantly affect
bloating.  Tank sizes varied from 800-1,200 bu capacity.  For
the control fermentations, freshly prepared, 6.6% NaCl cover
brine was used.  Recycled brine was added to cucumbers without
dilution.  The salt concentrations varied from 10-12% NaCl.
The salt concentration declined as the brine equilibrated with
the water in the cucumbers.  In all tanks dry salt was added
to the tanks as required to maintain the salt concentration of
6.6% (25° S) during the fermentation.  At the end of the
fermentation period, salt was added according to the usual
schedule to slowly raise the salt to 11%.

     Tanks were filled in groups of three consisting of a
control tank, a tank in which heated brine was used and a tank
into which chemically treated brine was added.  During the 1975
season, nine complete groups of tanks were fermented to give
nine replicates per treatment.  During the 1976 season, eight
groups of tanks were fermented.  In this instance, the number
of groups was limited by the amount of the second cycle
chemically treated brine available.
                               11

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            Fresh Salt Brine
                  1
      1974 1st Cycle Fermentation
                   I
         1st Cycle  Spent Brine
 Pasteurization
    Treatment
 1975 2nd Cycle
  Fermentation
       i
   2nd Cycle
  Spent Brine
       1
 Pasteurization
    Treatment
 1976 3rd Cycle
  Fermentation
   3rd Cycle
  Spent Brine
  Chemical
  Treatment
1975 2nd  Cycle
 Fermentation
     I
  2nd  Cycle
 Spent Brine

      t
  Chemical
  Treatment
                             I
1976 3rd  Cycle
 Fermentation
  3rd  Cycle
 Spent Brine
1975 Control
Fermentation
     I
  1st Cycle
 Spent Brine
1976 Control
Fermentation
                                                          i
  1st Cycle
 Spent Brine
Figure 3.   General  design of the commercial  evaluation
             of brine recycling.
                                12

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     When groups of tanks were filled, 1,000-lb tote boxes
of cucumbers were dumped alternately into the tanks to assure
similar cucumbers were put in each tank.  Fresh cucumber samples
were evaluated for mechanical damage, carpel separation, loose
seed cavities, and other defects.

     Recycled brine was sampled in the spring before and after
treatment prior to use in the fermentations.  These samples
were used for salt, pH, acid, reducing sugar, suspended solids,
BOD, COD, Kjeldahl nitrogen and mineral analyses.  Addition
of salt was recorded throughout the fermentation.  Brine from
each tank was sampled for the same analyses listed above in
the winter when cucumbers were removed for processing.  Two
5-gal pails of cucumbers were taken from each tank for analysis
of nitrogen and mineral components.  One pail of cucumbers was
desalted in the laboratory with tap water.  A 10 kg sample of
salt stock and desalted cucumbers was diced with a vegetable
dicer and homogenized without addition of water with a Bronwill
Polytron homogenizer.  A 1 liter sample of each homogenate was
kept for analysis.

     The brine and cucumber samples were refrigerated when they
were collected in the tank yard.  The samples were frozen in
the laboratory.  For mineral and Kjeldahl nitrogen analyses,
the samples were held from 1-6 months prior to analyses.  Brine
samples for determination of BOD, COD, suspended solids,
titratable acidity, and NaCl were refrigerated.  The BOD was
run within 3 days of receipt of the samples.  The other
analyses were completed within 1 week.

     To evaluate the commercial quality of salt stock produced
with recycled brine, samples of 200 cucumbers were taken from
each tank.  They were classified into three categories by tank
yard personnel.  Good cucumbers were those with no internal
defects.  Commercially acceptable cucumbers included the good
cucumbers plus those with slight bloater or honeycomb defects
which could be sliced into dill chips with only small losses.
Commercially unacceptable stock had severe bloater or honeycomb
defects which made it suitable only for less valuable relish
stock.  The texture of desalted cucumbers was evaluated by a
puncture test with a 5/16 inch diameter probe.  This was
similar to the test developed by Bell and Etchells (11)  using
a Magnus-Taylor manual pressure tester.  The test was done
with an Instron Universal testing instrument (Instron Corp.,
Canton, MA).  The force required to puncture the cucumber was
determined by running a 5/16 inch diameter probe vertically
into the fruit at a speed of 50 cm/min.  The recorder was set
to record 20 kg pressure full scale.  Twenty cucumbers from
each tank were punctured.  The average peak force required to
puncture the cucumbers was calculated.
                               13

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     A taste panel evaluation of the final product made from
2nd and 3rd cycle stock and the corresponding control stock was
conducted.  Hamburger dill chips from the 2nd and 3rd cycles
were prepared by desalting the stock and using commercial
procedures of Vlasic Foods, Inc.

     The difference test score sheet (Figure 4)  was used for
the taste test evaluation  (12)  ("Reference Guidebook for
Sensory Testing," 3rd edition, 1966, Continental Can Co., Inc.,
Chicago, IL).  The score sheet is divided into three parts.
In the first part the control sample is evaluated for flavor,
texture and overall quality in an "excellent" to "poor" rating.
In the second part the judges compare the heat and chemical
treatments with the control on the scale of 0 (no difference)
to 5 (very large difference).  Part III compares overall
quality of the treatments with the control product on the scale
of 1 (much worse) to 4 (same) to 7  (much better).  The data
were analyzed using analysis of variance procedures.

     During the fermentation of the cucumbers, the brines were
analyzed for salt, pH and titratable acidity.  These were the
normal analyses performed in commercial operations.  Salt was
measured using a hydrometer to determine the percent saturation
of a salt solution.  In addition, salt was measured by
titrating brine samples with 0.171 N AgNOj.  The results were
expressed as degrees salometer.  A 1% change in salt concen-
tration is approximately equivalent to a change of 4° salometer.
The pH was measured with a glass electrode.  Titratable acidity
was determined by titration of a brine sample with NaOH to
phenolphthalein end point or pH 8.  Results were expressed as
percent lactic acid.

     Bloater damage was evaluated on each commercial tank of
cucumbers according to the criteria used in standard commercial
operations at Vlasic.  The tanks were emptied from October to
March.  The salt stock was desalted to approximately 4% NaCl
according to the commercial practice of Vlasic Foods, Inc. and
processed into final products, primarily hamburger dill chips.

     Brine was analyzed for BOD  (13), COD  (14), and suspended
solids  (15) using standard methods.  Reducing sugar was
determined with the dinitrosalicylic acid reagent  (16).
Nitrogen was measured using a microKjeldahl procedure (17).

     Brine and cucumber samples were assayed for lead, cadmium
and chromium by atomic absorption spectrophotometry.  A method
described by Sprague and Slavin  (18) and by Thomas et al.  (19)
was used with quantities of samples appropriate for cucumbers
and brine.  A 40 ml aliquot of brine or a 40 g sample of ground
cucumbers was added to a 250 ml Erlenmeyer flask together with
35 ml of concentrated nitric acid.  The sample was boiled
                               14

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                        DIFFERENCE TEST
                                       Name:
                                       Date:
  I. Sample K is the standard, or reference sample.  Please rate
     K below:
        Overall Quality of K
           (circle one)	

            Excellent
            Good
            Fair
            Poor
          Comments
     Flavor
 Texture
    Excellent
    Good
    Fair
    Poor
Excellent
Good
Fair
Poor
 II. Please compare the samples to K for texture and flavor
     differences.  Test samples may or may not be different
     from the reference sample.  Rate each sample according to
     this difference scale:
            Rating

              0
              1
              2
              3
              4
              5
Difference from Standard

    No difference
    Very slight difference
    Slight difference
    Moderate difference
    Large difference
    Very large difference
Test Sample
No.


Difference Ratings
Texture


Flavor


Comments
Texture


Flavor


III. Quality of samples compared to K:

                              Test Sample #

     Much worse               	
     Moderately worse         	
     Slightly worse           	
     Same                     	
     Slightly better          	
     Moderately better        	
     Much better
               Test Sample #
 Figure 4.  Score sheet for taste panel evaluation of hamburger
            dill chips.
                               15

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gently on a hotplate for approximately 3 hr until it turned
clear and had reached a volume of 10 ml.  The sample was then
cooled to room temperature and the pH adjusted to 3.0 with
ammonium hydroxide.  After cooling again to room temperature,
5.0 ml of 2% ammonium pyrilidine dithiocarbamate was added.
The flask was swirled and then allowed to stand for 10 min.
Then 10 or 20 ml of methyl isobutyl ketone (MIBK) was added.
The solution was shaken for 2 min and allowed to stand until it
became clear.  The MIBK phase was then removed and analyzed.

     Mercury analysis was based on the method developed by
Hatch and Ott (20).  In determining mercury levels, 40 ml of
brine or 40 g of ground cucumber was transferred to a 250 ml
Erlenmeyer flask and dried on a hotplate at 71°C for approx-
imately 20 hr or until the sample appeared moisture free.  The
sample was cooled to -18°C, then 20 ml concentrated nitric acid
was added and the sample was allowed to stand for 1 hr at room
temperature.  After heating on a steam bath for 4 hr, 15 ml of
concentrated sulfuric acid was added and heating was continued
until the brown nitrous fumes were driven off (approximately
1 1/2 hr) .  The sample was cooled on ice and 15 ml of KMnC>4  (5%
w/v) was added.  The sample was heated on a steam bath for 30
min to complete the sample preparation.  When the solution was
cooled to room temperature, the mercury concentration was
measured with a Coleman model MAS-50 mercury analyzer.

     Phosphorus/ calcium, magnesium, manganese, iron, copper,
boron, zinc, and aluminum levels were determined by atomic
emission spectroscopy (21).  Brine or ground cucumber samples
were dried in porcelain crucibles on a hotplate at <100°C over
a 32-hr period.  The dried samples were ashed in a muffle
furnace at 400°C for 7 hr.  Samples were analyzed by the
Michigan State University Leaf Analysis Laboratory.

SPECIAL STUDIES

     Certain studies were done separate from commercial brine
recycling to evaluate potential difficulties with recycling.
These included an evaluation of pesticide buildup with multiple
reuse of brine, determination of the temperature and pH
stability of fungal pectinases which are most likely to occur
in  fermentation brines and a measurement of lysinoalanine in
chemically treated brine.
                               16

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Effect of Brine Recycling on the Buildup of Pesticide Residues

     Selected cucumber plots were treated with twice the
recommended dosage of pesticides at half of the recommended
intervals before harvest.  The plants were treated with
Terrachlor Super X (pentachloroaniline), Demesan  (chloroneb),
parathion, Thiodan (endosulfan), and Sevin (carbaryl).  Demesan
was not applied in 1977 because it was dropped from consider-
ation for use on cucumbers.

     The cucumbers were shipped to Michigan State University
and brined without sizing or washing.  Approximately 110 Ib
of cucumbers were placed in each of seven tanks and covered
with brine to give a pack-out ratio of 57:43 by weight of
cucumbers to brine.  For the first cycle all seven tanks of
fresh cucumbers were covered with fresh 6.6% NaCl solution.
In subsequent cycles the control tank was covered with fresh
brine, three tanks were covered with heat treated brine from
the previous cycle, and three tanks were covered with chemically
treated brine.

     The fermentations were carried out indoors under ultra-
violet lights to retard the growth of film yeasts.  Dry food-
grade salt was added to the tanks as required to maintain the
salt concentration at 6.6%.

     At the end of the fermentation, usually 3-4 weeks, the
cucumbers were removed from the tanks, the brines were treated
and used as a cover brine on the next batch of fresh cucumbers.

     Samples were taken for pesticide analysis of fresh washed
and unwashed cucumbers, brined cucumbers, and brine before and
after heat or chemical treatment.  The cucumbers for the
washed sample were sprayed with tap water from a spray nozzle
for 2 min.  The water flow rate was 10 kg/min.  For fresh and
brined cucumber samples, 10 kg of cucumbers were diced on a
vegetable slicer and then homogenized with a Bronwill Polytron
tissue grinder.

     Brines and cucumbers were analyzed by extraction of
pesticide materials followed by gas-liquid chromatography.
Solvents used in the analysis were methylene chloride, acetone,
ethyl acetate, and petroleum ether  (boiling range 30-60°C).
These solvents were obtained commercially, distilled in glass,
from Burdick and Jackson Laboratories, Inc., Muskegon, Michigan
49422.  Chloroneb, PCNB,pentachloroaniline (PCA), parathion,
paraoxon, Bravo®, Dathar^, Difolatari-^, endosulfan I,
endosulfan II, endosulfan sulfate, and carbaryl were obtained
from Pesticide Reference Standards Section, Chemistry Branch,
Registration Division, room 5175, South Agriculture Building,
Environmental Protection Agency, Washington, DC  20460.
                               17

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Bio Beads S-X3-200-400 mesh, control no. 13127, catalog no.
154-2750, were purchased from Bio-Rad Laboratories, Richmond,
California  94804.

     Two gas chromatographs were used for the analysis.  The
instruments, columns and operating conditions were as follows:

(1) Perkin Elmer model 910 equipped with nitrogen-phosphorus
    detector (NPD).

    Columns -1.8mx4mmid glass columns packed with 10%
    OV-101 and 10% OV-101 + 15% OV-210  (1 + 1), both on
    Chromosorb W-HP, 80-100 mesh, conditioned 48 hr at
    250 °C with ca.  20 ml/min N2 flow.

    Operating conditions - temperatures (°C) - column
    210, injector 230, detector 250; nitrogen carrier gas,
    60 ml/min;  NPD bead temperature - 10 turn pot setting
    630 or to give approximately 50% full scale deflection
    (FSD) for 8 ng carbaryl on 1 mv recorder, but not to
    exceed a dull red glow of detector bead.  Detector
    gas flows - hydrogen 3 ml/min, air 38 ml/min.
    Nitrogen and phosphorus containing compounds can
    both be determined under the above conditions,
    however, the response for phosphorus is extremely
    high.  To circumvent making large dilutions, the
    phosphorus compounds were quantitated with a flame
    photometric detector.

(2) Tracor model MT 222 equipped with linearized 63Ni
    electron capture detector  (ECD) and flame photometric
    detector (FPD) connected in parallel with column effluent
    splitter.

    Column -1.8mx4mmid glass column packed with
    10% OV-101 on Chromosorb W-HP, 80-100 mesh
    conditioned for 48 hr at 250°C with ca. 20 ml/
    min N2 flow.

    Operating conditions - temperature  (°C) - column
    200, injector 230, detectors 350  (ECD), 220  (FPD).
    Argon carrier gas - 60 ml/min, split at column
    exit with 1 part to ECD and 19 parts to FPD.
    Purge gas to ECD - 70 ml/min argon-methane  (95 +
    5).  ECD attenuation set to give 40-60% FSD for
    1 ng injected heptachlor epoxide with signal fed
    to 1 mv recorder.  FPD operating conditions -
    hydrogen 50 ml/min, air 80 ml/min, electrometer
    settings so that 2 ng of injected parathion gives
    40-60% FSD into a 1 mv recorder.
                               18

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     Gel permeation chromatography was performed with a GPC
Autoprep 1001 (Analytical Bio Chemistry Laboratories, Inc.,
Columbia, Missouri  65201) equipped with a 2.5 x 50 cm column
packed with Bio Beads S-X3.  Methylene chloride at a 5 ml/min
flow rate was used as the solvent.  Usual practice was to
discard the effluent for 29 min and then to collect the
sample for 24 min.  However, dump and collection times may vary
depending upon flow rate, column packing, etc.  Each column
and instrument should be calibrated with the compounds of
interest before analysis of samples is begun.

     Samples were concentrated with a 500 ml Kuderna-Danish
concentrator fitted with a three-ball Snyder column (Kontes
Glass Co., K-570000 or equivalent).  The lower joint of the
concentrator must fit a $19/22 of a 10 ml Mills-type graduated
tube (Kontes Glass Co., K-570050 or equivalent).

     Samples were blended with a Sorvall blender equipped with
a 500 ml, stainless steel cup (Ivan Sorvall, Inc., Newton,
CT  06470 or equivalent).

     Brine solutions were shaken before sampling to obtain an
even distribution of any settled material.  Raw and brined
cucumbers were chopped in a food chopper to give a homogenous
mass.  Samples were extracted according to the procedure of
Luke et al. (22).  Brine  (40 g)  was weighed into a 125 ml,
glass stoppered Erlenmeyer flask, 80 ml of acetone was added,
and the sample was thoroughly mixed.  For raw or brined
cucumbers a 100 g chopped sample was added to the blender cup,
200 ml acetone was added and the sample was blended at high
speed for 2 min.   The slurry was centrifuged and the super-
natant decanted through a glass wool plug into a 300 ml glass
stoppered Erlenmeyer flask.  Alternately, the sample could be
filtered with suction through a 12 cm Buchner funnel fitted
with sharkskin filter paper.

     Eighty ml of extracts, whether from brine or cucumbers,
was transferred to a 1 liter separatory funnel.  One hundred ml
each of petroleum ether and methylene chloride were added.
The funnel was shaken vigorously for 1 min, the layers were
allowed to separate and the lower aqueous phase was drained
into a 250 ml separatory funnel containing 7 g NaCl.  This
funnel was shaken for 30 sec to dissolve most of the NaCl.
The upper phase from the 1 liter separatory funnel was passed
through a 5 cm column of anhydrous granular Na^SC^ into a 500 ml
Kuderna-Danish concentrator fitted with a 10 ml graduated tube.
The 1 liter separatory funnel was rinsed with 100 ml methylene
chloride.  This solvent was then added to the 250 ml separatory
funnel to re-extract the aqueous phase.  The funnel was shaken
for 1 min and the lower methylene chloride phase was drained
through the Na2SC>4 column into the Kuderna-Danish concentrator.
                               19

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The aqeous phase was extracted with another 100 ml portion of
methylene chloride.  After extraction, this solution was also
dried and added to the concentrator.  The combined methylene
chloride extracts were evaporated to about 2 ml with the aid
of a three-ball Snyder column.  It was necessary to take care
to prevent solvent from boiling into the column at the
beginning of the evaporation.

     The concentrated extract was diluted to 7.0 ml with
methylene chloride in the graduated tube.  The sample (5.0 ml)
was loaded into the sample loop of the Autoprep 1001 and
chromatographed by dumping 29 min (145 ml) and collecting
for 24 min (120 ml).  "Cleaned up" extract was collected and
concentrated in the Kuderna-Danish concentrator to approxi-
mately 2 ml.   The tube containing sample was removed from
the concentrator.  The sample was evaporated to dryness with
a slight current of air or nitrogen and minimal heat.  The
dried sample was dissolved in 3.0 ml ethyl acetate for initial
injection on the gas chromatograph equipped with the NPD for
quantitation of carbaryl.  Appropriate dilutions were made for
injection on the ECD/FPD equipped chromatograph.

     Sample represented in the final ethyl acetate extracts
was calculated as follows:

     Cucumbers:  100 g/(200 ml + 94 ml - 10 ml) x 5 ml/7 ml;
     where 94 ml was the volume of water present in the
     cucumbers, 10 ml was the water-acetone contraction
     factor and 5 ml/7 ml was the quantity of sample
     used for GPC cleanup.

     Brine:  40 g/80 ml + 34 ml - 2.5 ml x 5 ml/7 ml;
     where 34 ml was the volume of water in the brine
     sample containing 15% solids and 2.5 ml was the
     water-acetone contraction factor.

Pectinase Inactivation

     The possibility for introduction of pectinases into spent
brines, either from the previous fermentation or during periods
of brine storage, was a major concern in the development of
recycling procedures.  If these enzymes were not properly
controlled, serious softening of cucumber salt stock could
result.

     Etchells et al. (23) did a survey of the fungi present on
cucumber fruits and flowers.  They identified the most common
molds and found that all of the species investigated produced
pectinase activity.  It was their opinion that enzymes from
the dominant molds would be the usual cause of commercial
cucumber softening.
                               20

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     Pectinase was produced by growing each of the organisms
studied on the following medium  (24) :  NI^NC^  (1 g/1) , MgSC>4
(0.3 g/1), yeast extract  (Difco)  (0.1 g/1), KH2P04  (0.01 M) and
citrus pectin  (Sigma)  (5.0 g/1).  The pH was adjusted to 6.4
with 4 N NaOH prior to sterilization of the medium.  Fernbach
flasks containing 1 liter of media were inoculated with
appropriate organisms and shaken  at  110 rpm on a rotary shaker.
Cultures were grown at room temperature (21-24°C) until enzyme
activity reached its maximum level.

     Fungal cells were removed from  the medium by centrifugation
for 5 min at 12,000 x g.  The supernatant was filtered through
a 0.45 y pore diameter Millipore  filter.  The cell free fil-
trate, containing pectinase activity, was concentrated with an
Amicon model 402 ultrafiltration  cell using a UM-10 membrane
with a molecular weight cutoff of approximately 10,000 daltons.

     For thermal inactivation studies pectinase concentrates
were added to brine solutions such that during inactivation 12%
NaCl, 0.6% lactic acid and 0.1% Ca++ ion were present in the
brine.  The brine pH was adjusted to 3.3 to 4.7 depending upon
the particular experiment.  Brines were heated for specific
time intervals in 4.5 mm I.D. test tubes sealed with paraffin
and cooled in an ice bath.  The enzyme activity remaining was
measured by the rate of viscosity change in a 1.0% pectin
solution in pH 4.0, .015 M lactate buffer.  The time required
for 90% inactivation of enzyme activity (D-value) was deter-
mined from the linear portion of  a plot of the logarithm of
the percent activity remaining as a  function of time.

     Inactivation of pectinase at high pH was done in simulated
spent brines in which Ca   ion was omitted to avoid formation
of a precipitate.  Glycine (0.02  M) was added to provide buffer
capacity at high pH in the brine.  It was necessary to flush
with nitrogen gas and tightly close test tubes with corks or
rubber stoppers to prevent the brine pH from dropping during
incubation.  The pH drop was apparently a result of CC>2
absorption during incubation.

     The pH of the high pH brines was adjusted to 4.0 with 1 N
HC1 prior to measuring the remaining enzyme activity.  Activity
was determined as described above.  The inactivation of pecti-
nases at high pH did not follow first order reaction kinetics.
As a result, D-values could not be determined.
                               21

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     Enzyme containing brines for inactivation experiments
in commercial spent brines were prepared by dialyzing enzyme
concentrate in a large excess of spent brine.  This was done
to prevent dilution of brine constituents by the enzyme
solution.

Lysinqalanine Content of Chemically Treated Brine

     Formation of lysinoalanine by base treatment of protein
had been reported by DeGroot and Slump (26). Woodward and Short
(27)  had reported that free lysinoalanine was toxic to rats.
Though cucumber brines are low in protein, the high pH
required for chemical treatment provided the potential for
formation of this compound.

     To determine whether significant amounts of lysinoalanine
could be formed by chemical treatment a first cycle, commercial
spent sample brine was used.  The brine was adjusted to pH 11.4
and held at room temperature (23°C) for 8 days.  The clear
supernatant brine was removed and the pH adjusted to 4.6 with
glacial acetic acid.  Samples of untreated and base treated
brine were dialyzed against water to remove salt and low
molecular weight components.  The dialyzed samples were
concentrated on a flash evaporator with the water bath heated
to 40°C.  Aliquots of the concentrates were hydrolyzed at
100°C with 6 N HC1.  The hydrolysate was chromatographed on
a Beckman, 120C amino acid analyzer using the column for
basic amino acid.  Authentic lysinoalanine was chromatographed
to verify the position on the chromatogram and for quantitative
standardization of the column.
                               22

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                           SECTION 5

                    RESULTS AND DISCUSSION

CHARACTERISTICS OF RECYCLED BRINES

     Brine samples were collected before and after heat and
chemical treatment for the first cycle brines in 1975 and for
second cycle brines in 1976.  Tables 1 and 2 show the mean
values for the results of the analysis done on these brine
samples.  The data were analyzed by an analysis of variance
procedure to determine the changes which occur as a result of
heat or chemical treatment.

     There were no significant differences between the untreated
brines used for the first cycle heat treatment and chemical
treatment in 1975.  This was expected since these brines were
randomly selected from the tank yard and assigned to either the
heat or chemical treatment.  A more limited analysis of brine
samples was done previously (8).  The levels observed in this
project for the mineral components of the brines were similar
to those obtained in the earlier study.

     Heat treatment caused few changes in the brine.  The pH
and titratable acidity were increased and decreased, respective-
ly.  NaOH was added after heat treatment to effect those
changes.  There was approximately a doubling of the aluminum
level.

     Chemical treatment caused a number of changes in the
brines.  The pH and titratable acidity changed as expected.
Reducing sugars declined under the alkaline conditions.
Formation and removal of a precipitate reduced the suspended
solids.  BOD and COD increased as a result of vinegar addition
for pH adjustment.  Several of the mineral components of brine,
including P, Ca, Mg and Fe, declined as a result of chemical
treatment.  It was found previously (8) that the precipitate
formed consisted primarily of mineral material.  Aluminum was
the only mineral component to show an increase.
                               23

-------
 TABLE 1.  CHARACTERISTICS OF 1ST CYCLE SPENT BRINE  IN  THE
           SPRING OF 1975 BEFORE AND AFTER HEAT OR CHEMICAL
           TREATMENT.  THE TREATED BRINES WERE USED  AS
           COVER BRINES FOR 2ND CYCLE FERMENTATIONS
Parameter
Acid % lactic
Salt %
pH
Sugar mg/1
Suspended solids rag/1
BOD mg/1
COD mg/1
Kjeldahl N mg/1
Cd mg/1
Hg mg/1
P mg/1
Ca mg/1
Mg mg/1
Mn mg/1
Fe mg/1
Cu mg/1
B mg/1
Zn mg/1
Al mg/1
Cr mg/1
Heat treatment
Untreated
0.45 B*
13.1
3.58 A
405 B
254 B
10300 b
14200 b
594
.015
0
130 B
1010 B
141 B
2.7
6.2 B
1.6
1.4
4.9
5.3 a
.003
Treated
0.17
12.5
4.63
272
204
9700
12800
507
.038
0
111
978
130
2.5
6.7
1.6
1.3
4.3
10.7
.003
A

B
AB
B
b
b



B
B
B

B



b

Chemical
Untreated
0.47
13.2
3.53
392
231
10600
14000
564
.093
0
116
1020
130
3.4
7.2
2.0
1.4
5.0
6.1
.003
B

A
B
B
b
b



B
B
B

B



a

treatment
Treated
0.23
12.7
4.80
190
55
14000
17200
523
.058
0
26
714
12
1.9
2.5
1.3
1.2
3.2
11.6
.001
A

B
A
A
a
a



A
A
A

A



b

*Samples with different letters are significantly  different.
 With upper case letters, differences are significant  at the  1%
 level.  With lower case letters, differences  are  significant
 at the 5% level.
                               24

-------
TABLE 2.  CHARACTERISTICS OF 2ND CYCLE  SPENT  BRINE  IN THE
          SPRING OF 1976 BEFORE AND AFTER  HEAT OR CHEMICAL
          TREATMENT.  THE TREATED BRINES WERE USED  AS
          COVER BRINES FOR  3RD CYCLE  FERMENTATIONS
Parameter
Acid % lactic
Salt %
pH
Sugar mg/1
Suspended solids mg/1
BOD mg/1
COD mg/1
Kjeldahl N mg/1
Cd mg/1
Hg mg/1
P mg/1
Ca mg/1
Mg mg/1
Mn mg/1
Fe mg/1
Cu mg/1
B mg/1
Zn mg/1
Al mg/1
Heat treatment
Untreated
0.62 B*
11.4
3.52 C
263 B
298
12500 b
18600 b
583
0.013
0.003
139 A
1042 A
173 A
1.4
9.3
1.4
1.4
4.8
8.3 A
Treated
0.21 A
11.8
4.55 A
222 B
206
14000 ab
19300 b
621
0.012
0.003
128 AB
1123 A
183 A
1.7
11.2
1.6
1.6
5.6
12.3 A
Chemical treatment
Untreated
0.63 B
11.0
3.66 B
258 B
158
14500 ab
19500 b
601
0.012
0.003
100 B
960 A
140 B
1.0
17.5
1.0
1.3
4.2
7.1 A
Treated
0.26
10.8
4.95
115
87
17400
22700
507
0.015
0.003
11.0
652
5
0.6
4.9
1.6
1.5
3.6
31.2
A

A
A

a
a



C
B
C





B
*Samples with different letters  are  significantly different.
 With upper case letters, differences  are  significant at the
 1% level.  With lower case letters, differences  are  signi-
 ficant at the 5% level.
                               25

-------
    In 1976 the second cycle spent brines were treated prior
to the third cycle fermentations.  Prior to treatment, a few
differences were observed between the brines which had been
heat treated and chemically treated the previous year.  The
pH of the chemical treatment brine was 3.66 compared to 3.52
for the heat treatment brine.  This difference was significant
at the 1% level.  It is thought to be the result of vinegar
addition which caused an increase in buffer capacity in the
chemical treatment brine.  Phosphorus and magnesium were also
reduced in the chemical treatment brines.

    Heat treatment of the second cycle brine resulted in no
significant change in brine composition except for the
increase in pH and decrease in titratable acidity.  Chemical
treatment caused significant changes in the same components
and in the same directions in 1976 and 1975 with the exception
of BOD and Fe.  In those two instances there were differences
in the same directions as were observed in 1975.  However,
the differences were not significant at the 5% level.

    Comparison of the heat treated first and second cycle
brines showed an increase in BOD from 9700 mg/1 to 14,000 mg/1
and COD from 12,800 mg/1 to 19,300 mg/1.  In addition, there
were small increases in the concentration of P, Ca and Mg.
For chemical treatment this first and second cycle comparison
showed that BOD and COD increased.  Though the levels of P,
Ca and Mg were lower in treated brines, the levels in 1975
were actually slightly higher than in 1976.  This may have been
a result of more efficient removal of precipitate in 1976.

    The results of these experiments showed that heat treatment
had no effect upon the brine constituents, except for aluminum
and the pH and titratable acidity, which were intentionally
altered.  Chemical treatment results in a decline of many, but
not all, of the mineral constituents of brine.  The BOD and
COD are increased as a result of vinegar addition.  Aluminum
levels increased after completion of brine treatment with both
the chemical and heat techniques.  It may be that the food
grade NaOH, which was used in both procedures, contains some
aluminum as an impurity.

    Third cycle brines were removed from cucumber fermentation
tanks after the third cycle fermentations.  The composition of
the third cycle brines is given in Tables 5 and 6.  The
concentrations of mineral constituents were similar to the
concentrations before treatment of the first and second cycle
brines.  Since the project was concluded at the end of the
third cycle of fermentation, the third cycle brines were not
treated.
                              26

-------
 CHARACTERISTICS OF CUCUMBERS AND FERMENTATIONS

     Tables 3 and 4 summarize the numbers of tanks operated in
this project and the amount of cucumbers and brines used.  The
control tanks were covered with 6.6% (25° S) NaCl brine at the
beginning of cucumber fermentation.  This was the usual com-
mercial practice for using fresh brines.  Recycled brines were
used at whatever salt concentration was present after brine
treatment.  This averaged about 12.5% in 1975 and 11% in 1976.
The ability to use a high salt cover brine is a necessary part
of successful recycling, since dilution of brine to 6.6% would
require discharge of a large fraction of spent brine due to
the excess volume generated.

     The cover brine was diluted as it equilibrated with the
water in the cucumbers.  To maintain the salt concentration at
6.6% during fermentation, dry salt was added to the tank head-
boards.  These tanks were fermented as nonpurged, natural
fermentations.  This was the usual fermentation procedure at
the time this project was initiated.  Recently Costilow et al.
(28) reported on a nitrogen-purging procedure which was
designed to remove C02 from natural fermentations.  The
technique was shown to significantly reduce bloaters during
cucumber fermentations.  The prospect is that a significant
proportion of commercial fermentations will be using this tech-
nique in the immediate future.  Vlasic has used the purging
technique in combination with recycled brine without difficulty
in fermentation tanks which were not part of this project.  No
variations in brine composition would be anticipated as a
result of using purging, since the only effect of the technique
is to circulate the brine during the fermentation period in
such a way that COo generated by the microorganisms and the
fruit can be efficiently removed.
                               27

-------
TABLE 3.  SUMMARY  DATA ON  FERMENTATION TANKS, 1975

Number of tanks
Bushels of cucumbers
Gallons of brine
Gallons of brine/bushel
Pack-out ratio
TABLE 4. SUMMARY DATA ON

Number of tanks
Bushels of cucumbers
Gallons of brine
Gallons of brine/bushel
Pack-out ratio
Control
10
7660
34321
4.48
57:43
Heat
treatment
11
8668
31418
3.62
62:38
FERMENTATION TANKS,
Control
8
5727
19832
3.46
63:37
Heat
treatment
8
5604
20780
3.71
62:38
Chemical
treatment
10
7388
30881
4.18
59:41
1976
Chemical
treatment
8
5727
19960
3.49
63:37
                              28

-------
     Tables 5 and 6 show the composition of spent brines at the
time cucumbers were removed from the fermentation tanks.  The
control brines in 1975 and 1976 were first cycle spent brines.
The tanks which contained heat and chemically treated brines
were second cycle fermentations in 1975 and third cycle fermen-
tations in 1976.  When second cycle brines were compared to
the 1975 first cycle control brines, the only significant
differences were a higher final pH in the recycled brine tanks
and a higher aluminum level in the heat treated brine than in
the control.  The heat and chemical treated brines differed
from each other in suspended solids, but neither was signifi-
cantly different from the control.

     Comparison of third cycle brines with the 1976 first cycle
control brines showed an increase in pH, reducing sugar, BOD
and COD in the chemically treated brines.  The heat treated
brines differed from the control only in pH, BOD and COD.  The
heat treatment brines had a higher Ca and Mg content than the
chemical treatment.  However, the control tanks showed inter-
mediate levels which were not significantly different from
either the heat or chemical treatment brines.

     The rise in final pH in recycled brine tanks was expected
because the buffer capacity of the recycled brines would
retard the drop in pH during fermentation.  BOD and COD levels
in brines were also expected to increase because of the carry-
over of organic matter when brines were recycled.  None of the
changes affected the course of fermentation to any practical
degree.  Observations of the progress of fermentations and the
tank yard operations with recycled brine showed that recycling
can be done without changing the tank yard operation during
the fermentation period.
                               29

-------
 TABLE 5.  ANALYSIS OF  SPENT  BRINE REMAINING IN THE FERMEN-
           TATION TANKS  IN THE  WINTER AFTER REMOVAL OF SALT STOCK
           CUCUMBERS FROM THE 2ND CYCLE (1975)  AND 3RD
           CYCLE  (1976) FERMENTATION TANKS

2nd cycle, 1975 data
Acid % lactic
Salt %
pH
Sugar mg/1
BOD mg/1
COD mg/1
Suspended solids mg/1
3rd cycle, 1976 data
Acid % lactic
Salt %
PH
Sugar mg/1
BOD mg/1
COD mg/1
Suspended solids mg/1
Control
0.67
9.6
3.36 A*
367
14700
17600
342 ab
0.57
8.6
3.55 a
186 A
12500 A
17000 A
552
Heat
treatment
0.68
10.1
3.48 B
415
14300
19200
553 b
0.74
9.7
3.65 b
176 A
19200 B
23900 b
379
Chemical
treatment
0.73
9.4
3.57
325
16300
20500
265
0.77
8.7
3.65
259
19600
23100
612


B



a


b
B
B
b

*Samples with different  letters are significantly different.
 With upper case letters,  differences are significant  at the
 1% level.  With lower case  letters, differences are signif-
 icant at the 5% level.
                               30

-------





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     Salt stock quality was evaluated for cucumbers fermented
in recycled brines.  Bloating has traditionally been a major
source of defects in fermented fruit.  Bloating results in sig-
nificant economic loss because seriously bloated fruit can only
be utilized in lower price relish products.  Tables 7 and 8 show
the results of bloater evaluations on second and third cycle
salt stock.  In both 1975 and 1976, the control was first cycle
salt stock.  No significant differences were found between
fruit fermented in recycled brines and the controls.  This
showed that bloater defects will neither increase nor decrease
as a consequence of recycling.  As stated above, if fermenta-
tions were properly purged, significant decreases in bloater
defects could be obtained.

   TABLE 7.  QUALITY OF 2ND CYCLE SALT STOCK CUCUMBERS
             FERMENTED IN RECYCLED BRINE, 1975*
Control Heat Chemical
treatment treatment
Good stock (%) 72.8 72.8
Commercially acceptable (%) 85.3 86.5
Commercially unacceptable (%) 14.7 13.5
73.0
86.5
13.5
*No significant differences among treatments.
TABLE 8. QUALITY OF 3RD CYCLE SALT STOCK CUCUMBERS
             FERMENTED IN RECYCLED BRINE, 1976*


Good stock (%)
Commercially acceptable (%)
Commercially unacceptable (%)
Control

52.1
84.9
15.1
Heat
treatment
46.0
80.2
19.9
Chemical
treatment
54.0
82.6
17.4
  *No significant differences among treatments.
                               32

-------
     In addition to bloater defects, the salt stock was
routinely evaluated for overall commercial acceptability at the
time of processing.  Cucumbers fermented in recycled brine were
judged to have normal texture, color and odor.  Instron pressure
test evaluation of the desalted stock from second cycle
fermentations showed no significant differences in the stock
from recycled brines.  The average pressure test was 17.5 Ib for
the control, 18.4 Ib for the stock from heat treated brine and
18.3 Ib for stock from chemically treated brine.  On the scale
proposed by Bell et al. (11), the fruit would be rated very
firm.

     Taste panel evaluations were done on hamburger dill chips
prepared from control salt stock and from salt stock from
second and third cycle fermentations.  Tables 9 and 10 show the
average scores for second and third cycle products and
corresponding control samples.  The last column shows the
cumulative averages.  The control samples were rated fair to
good when evaluated for flavor, texture and overall quality.

     Tables 11-15 show the results of analysis of variance of
the differences in texture, flavor and overall quality of
products prepared from control and recycled fermentations.
There were no significant differences between the control and
product prepared from salt stock fermented in either heat
treated or chemically treated, recycled brine.

     Based upon commercial experience, there does not appear
to be any significant deterioration or improvement in the
quality of products obtained from fermentation in recycled
brine.  There have been no complaints from either institutional
or individual consumers of these products which have been
related to use of recycled brine.
                               33

-------








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-------
TABLE 11.  ANALYSIS OF VARIANCE  TABLE  FOR TEXTURE
           DIFFERENCES OF HAMBURGER DILL CHIPS MADE
           FROM 2ND CYCLE SALT STOCK CUCUMBERS*
Source
Treatments
Groups
Treatments x groups
Judges
Judges x treatments
Judges x groups
Judges x groups x
treatments
df
1
6
6
8
8
48
48
ss
3.50
33.16
12.78
23.59
2.86
141.91
59.86
MS
3.50
5.53
2.13
2.95
0.36
2.96
1.25
F
1.64
2.60

1.00
0.12


*No significant differences
                          36

-------
 TABLE 12.  ANALYSIS OF VARIANCE TABLE  FOR FLAVOR
            DIFFERENCES OF HAMBURGER DILL  CHIPS  MADE
            FROM 2ND CYCLE SALT STOCK CUCUMBERS*
Source
Treatments
Groups
Treatments x groups
Judges
Judges x treatments
Judges x groups
Judges x groups x
treatments
df
1
6
6
8
8
48
48
ss
2.57
46.65
37.76
12.29
7.43
105.49
40.24
MS
2.57
7. 78
6.29
1.54
0.93
2.20
0.84
F
0.41
1.24

0.70
0.42


*No significant differences,
                            37

-------
  TABLE 13.  ANALYSIS OF VARIANCE TABLE FOR TEXTURE
             DIFFERENCES OF HAMBURGER DILL CHIPS
             MADE FROM 3RD CYCLE SALT STOCK CUCUMBERS
Source
Treatments
Groups
Treatment x groups
Judges
Judges x treatments
Judges x groups
Judges x groups x
treatments
df
1
4
4
10
10
40
40
ss
4.0
56.68
6.96
26.05
12.10
127.32
59.54
MS
4.0
14.17
1.74
2.61
1.21
3.18
1.49
F
2.30
8.14*

0.82
0.38


*Significantly different at p =  .05.
                           38

-------
  TABLE 14.  ANALYSIS OF VARIANCE TABLE FOR FLAVOR
             DIFFERENCES OF HAMBURGER DILL CHIPS
             MADE FROM 3RD CYCLE SALT STOCK CUCUMBERS*
   Source                     df     ss     MS     F


 Treatments                    1    20.08  20.08  9.21

 Groups                        4    33.81   8.45  3.88

 Treatments x groups           4     8.70   2.18

 Judges                       10    16.29   1.63  0.74

 Judges x treatments          10    15.02   1.50  0.68

 Judges x groups              40    87.99   2.20

 Judges x groups x
   treatments                 40    36.70


*No significant differences.
                           39

-------
 TABLE 15.  ANALYSIS OP VARIANCE TABLE FOR OVERALL
            QUALITY DIFFERENCES OF HAMBURGER DILL
            CHIPS MADE FROM 3RD CYCLE SALT STOCK
            CUCUMBERS*
Source
Treatments
Groups
Treatments x groups
Judges
Judges x treatments
Judges x groups
Judges x groups x
treatments
df
1
4
4
10
10
40
40
ss
6.64
0.86
9.68
22.70
13.26
126.94
64.92
MS
6.64
0.22
2.42
2.27
1.33
3.17
1.62
F
2.74
0.09

0.72
0.42


*No significant differences.
                           40

-------
     The concentrations of several minerals in the salt stock
and the desalted cucumbers were analyzed to determine the degree
of buildup of these materials which might result from recycling
brine.  Results of these analyses are summarized in Tables 16
and 17.  The second cycle salt stock showed no significant
differences in the mineral content compared to the control first
cycle salt stock except that at the 5% significance level, the
level of Al in the heat treatment salt stock was higher than
for the control or chemically treated samples.  The third cycle
samples from heat treated brine had higher levels of Mg, Mn
and Fe than the control.  There were no significant differences
between the samples from chemically treated brine and the
control samples.

     After desalting the salt stock cucumbers in the second
cycle samples, the only significant difference was a higher Mn
level in the samples from heat treated brine and a lower Al
level in samples from chemically treated brine compared to heat
treated brine samples.  The third cycle samples showed no
differences between the control and either recycled brine
technique.  The only difference significant at the 5% level was
a higher Mg concentration in samples from heat treated brines
compared to samples from chemically treated brines.  In several
instances there were lower levels of minerals in the desalted
cucumbers in 1975 compared to 1976 in all three groups of
samples.  This was attributed to the fact that in 1975 the
cucumbers were desalted to <1% NaCl.  In 1976 desalting was
done to bring the salt concentration down to 4%.  This was
closer to actual commercial practice.

     The results of the mineral analysis did not show any
buildup of elements to illegal or potentially toxic levels in
the spent brines (Table 6), the salt stock (Table 16) or the
desalted cucumbers (Table 17).  The Pb was below detectable
levels.  The lowest detectable level was estimated to be
approximately 0.05 ppm.  Samples were analyzed for chromium in
1975.  Most samples contained no detectable chromium.  As a
result, this analysis was not done in 1976.

     The data on the commercial quality of salt stock, on the
composition of spent brines before and after treatment, on the
course of fermentation and on the mineral components of
cucumbers before and after desalting led to the following
conclusions regarding the technological aspects of brine
recycling.

1. Major changes in tank yard practice were not required for
   brine recycling.

2. Fermentations proceeded normally.  The same criteria used to
   judge completion of fresh brine fermentations could be used
   to judge fermentations with recycled brine.

                               41

-------



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3. Treated brine which contained 12.5% NaCl could be used with-
   out dilution as a cover brine.   This yielded salt stock
   equivalent in quality to control tanks.

4. Changes in brine or cucumber composition as a result of
   recycling were small.  There were no significant buildups
   of toxic elements in brines or cucumbers which would
   indicate that recycling should not be done or that the
   number of brining cycles would need to be limited.

5. From a technical point of view, recycling the brine was
   practical and resulted in final products which were commer-
   cially equivalent to the controls.

ECONOMIC ANALYSIS OF BRINE RECYCLING

     The economics of brine recycling must be analyzed for each
particular case.  The costs and savings realized for this
project are summarized in Tables 18, 19 and 20.  Table 18 shows
the savings to be realized as a result of recycling brine.  Salt
which was reused represented a direct savings.  The calculation
was done on the basis of 12% salt in spent brine.  The savings
in treatment costs can be quite variable.  In this project waste
was treated in an on-site, aerated lagoon.   A cost of SC/lb BOD
treated was estimated.  No dollar cost could be placed upon the
fact that less salt was discharged as a result of recycling.
However, the plant had a stringent limit upon its discharge
of salt.  Reduction of NaCl levels in the effluent was
essential to continued operation of the plant.  A small cost
saving was realized as a result of reuse of water.

     The costs of the chemical treatment procedure are shown in
Table 19.  The labor cost was difficult to evaluate because of
the small scale of the chemical treatment operation.  Since the
time required was intermittent rather than continuous, as was
the case with the heat treatment procedure, the labor cost was
estimated to be 50% of the labor cost for pasteurization.  The
major costs were NaOH and vinegar.  In 1975 an average of 44 Ib
NaOH/1,000 gal of treated brine was required.  In 1976 this
increased to 52 Ib.  The vinegar use, calculated on the basis
of equivalent glacial acetic acid, also increased from 30 Ib/
1,000 gal in 1975 to 36 lb/1,000 gal in 1976.  These increases
may reflect an increase in buffer capacity of the second cycle
spent brine compared to first cycle brine.  The high cost of
chemicals resulted in a small net cost for chemical treatment.
No capital investment was required for the project.  However,
if chemical treatment were used as the sole brine treatment
procedure in a tank yard, the quantities required might make it
necessary to purchase storage tanks for the NaOH and acetic acid
along with metering equipment.
                               44

-------
  TABLE 18.  SAVINGS FROM RECYCLING 1,000 GAL OF  SPENT BRINE
        Item
             Dollars saved
  NaCl, 1.08 Ib/gal @ $20/ton
  BOD, 12,000 mg/1 at 8<=/lb
  Water, 47<=/l,000 gal
  Total savings
                  10.76
                   8.00
                    .45
                  19.21
 TABLE 19.  ECONOMIC EVALUATION OF CHEMICAL TREATMENT OF
            SPENT BRINES
       Item
  1975 Cost
($/l,000  gal)
   1976 Cost
($/l,000 gal)
Labor @ $4/hr
NaOH, 1975 19.SC/lb
      1976 17«/lb
Vinegar, 1975 $1.01/gal
         1976
   Total cost
   Total savings
   Net cost
    0.84
    8.58

   11.87

   21.29
   19.21
    2.08
     1.32
     8.84

    13.32

    23.48
    19.21
     4.27
                              45

-------
TABLE 20.  OPERATING COSTS AND SAVINGS USING HEAT
           TREATMENT FOR SPENT BRINES
Item
Labor @ $4/hr
Propane
Pumping
NaOH, 1975 19.SC/lb
1976 17«/lb
Total operating costs
Total savings
1975 Cost
($/l,000 gal)
1.67
1.23
0.71
2.76
6.37
19.21
1976 Cost
($/l,000 gal)
2.64
1.29
0.71
2.41
7.05
19.21
  Net savings based
   upon operating costs   12.84              12.16
                            46

-------
     The operating costs for heat treatment were considerably
lower than for chemical treatment.  A major cost was the
purchase of a $44,575 APV titanium alloy regenerative heat
exchanger with a brine treatment capacity of 50 gal/min.  This
first heat exchanger has been used for three brining seasons
without showing signs of major deterioration.  In 1976 the cost
of a new 50 gal/min capacity heat exchanger unit, with some
improvements and revisions of the first model, was approximately
$60,000.  The present cost of a basic turn-key unit with a
capacity of 50 gal/min with minimum instrumentation and
convenience features is $50,000.  A similar unit with a 35 gal/
min capacity costs approximately $40,000.

     An estimate of whether it is worth purchasing a heat
exchanger unit, considering the difference in operating costs
between chemical and heat treatment of brine, was made by doing
a calculation of the net present value (29) of the difference
in operating costs between chemical and heat treatments.
Such an estimate is highly dependent upon the assumptions used
for the calculation.  Each company considering brine recycling
must make the decision between treatments based upon their
particular circumstances.

     The results of the net present value calculation are shown
in Table 21.  The following assumptions were used in making the
calculation.

1. A heat exchanger unit with a treatment capacity of 35 gal/min
   will be purchased for $40,000.

2. The heat exchanger unit will be utilized at full capacity
   14 hr/day, 20 days/month for 3 months to treat 1.7 million
   gal of brine.

3. The interest rate will be 8.5%.

4. The difference between the operating costs of chemical and
   heat treatments will remain constant.

5. The maintenance cost of the brine recycling heat exchanger
   will be $500/yr.

6. The useful life of the heat exchanger unit will be 5 yr.

7. The value of the unit will be zero after 5 yr.

8. The capital cost of the chemical treatment will be zero.
                               47

-------
TABLE 21.  NET PRESENT VALUE CALCULATION FOR COMPARISON OF
           CHEMICAL TREATMENT WITH HEAT TREATMENT OF BRINE.
           THE DIFFERENCE BETWEEN TREATMENT COSTS PER
           1,000 GAL FOR CHEMICAL COMPARED TO HEAT
           TREATMENT OF BRINE WAS $16.43 IN 1976.  FOR 1.7
           MILLION GAL THIS IS A TOTAL DIFFERENCE IN
           TREATMENT COST OF 1700 x $16.55 = $27,931.
           WITH A MAINTENANCE COST OF $500/YR FOR THE
           UNIT THE NET DIFFERENCE IS $27,431.
   Years of use of the                    Present value of
   heat exchanger unit                    $27,431 at 8.5%
                                              interest
1
2
3
4
5
25,282
23,301
21,476
19,794
18,243
   Total present value of difference
    in costs between chemical and           	
    heat treatments                          108,096

   Present value of heat exchanger unit       40,000
   Net present value                          68,096
                             48

-------
     Based upon these assumptions, the present value of the
savings to be realized over a 5-yr period by using heat treat-
ment is over $108,000.  Since the cost of the heat exchanger
will be about $40,000, resulting in a net present value of over
$68,000, the heat treatment procedure would be the treatment of
choice.  This calculation was repeated making the same
assumptions as before except that only 50% as much brine was
treated.  In that case there was still a positive net present
value of over $13,000 when heat treatment was used.

     In summary, it appears that either chemical or heat treat-
ment can be utilized without making the cost of brine recycling
prohibitive.  It appears that in many situations, the heat
treatment procedure would be economically advantageous.
However, an analysis must be made for each particular situation
considering local economic and physical conditions.

POTENTIAL FOR REDUCTION OF WASTE

     Table 22 shows the raw waste loads for fresh pack, process
pack and pickle salting stations as published by EPA (30).
On April 16, 1976 the EPA published in the Federal Register (31)
a notice of effluent guidelines for existing sources and pre-
treatment standards for new and existing sources covering the
canned and preserved fruit and vegetable point source category.
Table 23 shows the effluent guidelines which must be met as of
July 1, 1977 using "best practical control technology currently
available" (BPCTCA).  The "best available technology economical-
ly achievable"  (BATEA) must be in place by July 1, 1983 to
meet the effluent limitations shown in Table 24.

     EPA has not developed limitations on total dissolved solids
(salt).  However, many states have placed limits on salt
discharge.  Therefore, in the design of waste reduction systems,
processors must consider reductions in salt as well as BOD,
TSS and flow rates.

     The problem of spent brine discharge from a tank yard must
be considered both in terms of direct BOD and TSS discharge
and its detrimental effects upon the waste treatment system.
If the spent brine is to be accomodated in an existing waste
treatment facility, it can cause several problems.  The
increased salt levels will reduce oxygen solubility, thus
lowering treatment efficiency, and the intermittent nature of
this high BOD, high salt wastewater will cause a shock effect
to most biological treatment systems.  The ultimate result will
be higher costs for the treatment operation.
                               49

-------
        TABLE 22.   RAW WASTE LOAD SUMMARY*
Item Flow gal/ton
1977 1983
Fresh pack 2051 1878
Process pack 2298 1481
Salting stations 253 77
BOD
1977
19.0
36.7
15.9
Ib/ton
1983
5.89
17.2
3.17
TSS
1977
3.82
6.54
0.83
Ib/ton
1983
1.91
1.57
0.43
*EPA 440/1-75/046, p. 197.   Development Document for Interim
 Final and Proposed Effluent Limitations Guidelines and New
 Source Performance Standard.
      TABLE 23.  EPA EFFLUENT GUIDELINES - 1977*
Item
Fresh pack
BOD
TSS
Process pack
BOD
TSS
Salt stations
BOD
TSS
Raw
Daily
maximum
1.22
2.19
1.45
2.63
0.18
0.33
(lb/1,000 Ib)
30 Day
average
0.75
1.54
0.92
1.91
0.12
0.25
Annual
average
0.53
0.99
0.68
1.28
0.09
0.18
   *Best Practical Control Technology Currently Available
    (BPCTCA).   Effective 7/1/77.
                               50

-------
      TABLE 24.  EPA EFFLUENT GUIDELINES - 1983*
Item
lb/1,000 Ib
Daily
maximum
30 Day
average
Annual
average
Fresh pack
BOD
TSS
medium
large
medium
large
0.639
0.639
1.139
0.639
0.461
0.461
0.606
0.461
0.213
0.213
0.429
0.213
Processed pack


TSS

medium
large
medium
large
0.652
0.652
1.208
0.652
0.511
0.511
0.784
0.511
0.313
0.313
0.643
0.313
Salt stations
BOD


medium
large
medium
large
0.084
0.084
0.163
0.084
0.072
0.072
0.125
0.072
0.054
0.054
0.113
0.054
*Best Available Technology Economically Achievable (BATEA)
 Effective 7/1/83.

 Medium - 2,000 to 10,000 tons/yr.

 Large - 10,000 tons or more/yr.
                           51

-------
     Table 25 shows the theoretical salt use based upon the
recommendations of Etchells and Moore (3)  for the fermentation
of cucumbers.  The amount of salt added after fermentation is
completed varies from company to company.   In most instances
the final level is 11% (45° S)  NaCl or higher.  The table shows
the amount of salt used at several final salt concentrations.
If brine recycling is not used, the salt discharge will be that
shown in the final column of the table.   If fermentation brines
could be 100% recycled, the salt in the spent brine would be
recovered and reused.

     The salt removed in the process water must still be dis-
charged with current technology.  Reduction of this source of
waste can be accomplished only by developing new fermentation
procedures which will allow storage at lower salt concentrations
and/or developing new economical procedures for concentrating
and reusing the dilute process water.

     The heat treatment procedure should result in recycling
approximately 95% of the spent brine with its organic and
inorganic waste load.  Removal of the precipitate layer formed
by chemical treatment will limit brine recovery to approximately
90%.  It must be pointed out that the brine recovery
accomplished in this project was achieved with tanks which were
not designed to recover the brine.  Improvement in this aspect
of recycling is certainly feasible if future tanks are
designed to maximize brine recovery.  For example, a conical
tank bottom would minimize the volume of brine containing
excessive dirt and debris and increase the percentage of brine
in a tank which could be readily pumped out.

     A salt balance study was done to determine the quantity of
salt which is lost from tank leakage and runover.  This study
showed a loss of 0.25 to 1.0 Ib of salt per bu of cucumbers.
The losses from leaky tanks can be partially controlled by
careful maintenance of the tanks.  However, development of
economical liners for wooden tanks or conversion to fiberglass
or other tank materials will probably be necessary for effective
control of this effluent.  Tank overflow during rainy periods
can only be stopped if fermentation technology is developed
which allows tanks to be covered during fermentation and storage,
The controlled fermentation process developed by Etchells et al.
(10) offers this possibility.
                                52

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     It appears that brine recycling is an economically
attractive operation for inclusion into process packers'  environ-
mental control systems.  However, effluent problems can not be
solved simply by introduction of brine recycling into the tank
yard operation.  Kirk  (2)  reported that spent brine accounted
for 8% of the total hydraulic load, 44% of the BOD and 59% of
the salt generated in a pickle salting station.  Even if
complete recycling of spent brine were achieved, it would be
necessary to greatly reduce tank leakage, tank overflow,
spillage and most importantly, desalting water  (process water)
in order to meet the 1983 limitations.  Therefore, a continuing
development of new technology must occur if the pickling
industry is to remain viable.  Effective application of brine
recycling technology will be one step in this process.

RESULTS OF SPECIAL STUDIES

Pesticide Distribution in Brine Recycling

     The study was performed to evaluate conditions representing
the extreme misuse of pesticides.  The cucumbers were raised
on a selected plot of land and were treated at twice the recom-
mended dosage with pesticides normally used by cucumber growers.
In addition, the cucumbers were harvested at half the post-
harvest interval recommended by the Environmental Protection
Agency.

     Methodology to determine the selected compounds simultan-
eously could not be found.  However, the procedure of Luke et al.
(22) extracts many more pesticides than are included in this
study and is rapid and easy to perform.  The decision was made
to expand this extraction into a suitable method for the
selected compounds.  Since the extract was too dirty for direct
quantitation using ECD and since several of the compounds are
not recovered through the PAM Florisil cleanup  (35), gel
permeation chromatography was investigated.

     Initially a 2.5 x 25 cm column packed with Bio Beads S-X3
and methylene chloride mobile solvent was tried.  The separation
between the sample extractives and the pesticides was borderline
at best and the decision was made to try a 50 cm column.
Separation was improved although some of the extractives had to
be collected to insure good recoveries of parathion and
paraoxon.  These extractives did not interfere with the GLC
quantitation by ECD.
                                54

-------
     The effect of direct evaporation of the GPC eluant of
methylene chloride from the collection beakers was compared to
evaporation using a Kuderna-Danish concentrator  (Table 26).
The recovery of pesticides from cucumbers, brines and with no
sample substrate is also shown in Table 26.  The unusually low
recoveries of Bravo® and Difolatan® in the Kuderna-Danish
evaporation compared to the higher recoveries through the
proposed method could not be explained.  The final evaporation
to dryness is critical.  The methylene chloride must be
removed; however, if the tube is allowed to remain under the
air or nitrogen jet after dryness, loss of residues can occur,
especially chloroneb, PCNB, PCA and Bravo .

     The cucumbers for pesticide distribution studies were
harvested, analyzed for residues and brined in seven 55-gal
drums.  At the end of each brining cycle, a portion of the
brining solution was removed from each of the seven drums and
saved for analysis as untreated brine.  The remainder of the
brines in drums 1, 3 and 4 were screened to remove large debris
and pasteurized at 175°C for 25 sec in a heat exchanger.
After treatment, the brines were pumped back into their
respective drums and the pH of the brine was adjusted with
NaOH just piror to reuse.

     The remainder of the brines in drums 5-7 were treated with
NaOH to raise the pH to 11.0 and allowed to stand for 48 hr.
The pH was then adjusted with acetic acid and the large debris
was screened out just prior to reuse.

     The treated brine in each drum was then reused with
another batch of pesticide treated cucumbers.  This process was
repeated for each cycle.  Three cycles were performed in 1975
and an additional three cycles were completed in 1976.  The
same brining solutions were used throughout the six cycles.

     The brine in drum no. 2 was carried through the six cycles
as a "control" and was neither pasteurized nor chemically
treated except to readjust the salt content or the pH.

     The effects of the pasteurization and chemical treatments
on the residues in the brining solutions can be seen in Tables
28-31.  Chloroneb®, Bravo®, Dacthal , Difolatan®, and paraoxon
showed no more than trace residues in any of the samples run
through the study and were not included in any of the tables
except Table 27, which gives the screening levels for each of
the pesticides.  Initial injections of sample extract equiva-
lent to 40 mg were necessary to attain these sensitivities.
Any peak less than 10% FSD was considered a trace and was not
quantitated.
                               55

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TABLE 27.  LOWEST LEVELS AT WHICH RESIDUES WERE
           QUANTITATED
Pesticide
Chloroneb
PCNB
PCA
Parathion
Paraoxon
Bravo
PPM
0.10
0.002
0.003
0.01
0.02
0.002
Pesticide
Dacthal
Difoltan
Endosulfan I
Endosulfan II
Endosulfan SO*
Carbaryl
PPM
0.004
0.02
0.01
0.01
0.02
0.05
                         57

-------
     Tables 28-29 indicate that carbaryl is the only one of the
selected pesticides that accumulates significantly if each
batch of raw cucumbers added to the tank would contain high
levels of residues (Table 28, cycles 1 and 2, "heat treated
brine" and "brined cucumbers").  The low residues on the raw
cucumbers used in cycles 3-5 were caused by heavy rains just
after the crop was sprayed with the pesticides.  These
unexpected low residues actually aided the study by showing
that if a batch of cucumbers with low residues is brined in the
tank, the levels in the brined cucumbers increased to slightly
more than in the raw cucumbers and the brine levels decreased
to almost trace amounts.  This was probably caused by the raw
cucumbers absorbing pesticides from the solutions during the
brining process (Table 28, cycle 3).  If several cycles of
cucumbers with high residues were brined in the same tank and
residues of carbaryl (or any of the pesticides tested) reached
very high levels, the brine could still be salvaged by one
chemical treatment.  As shown in Tables 28-31, the chemical
treatment procedure destroyed all detectable residues except
traces of PCNB and PCA in cycle no. 2.

Pectinase Inactivation

     Table 32 shows the list of organisms investigated.
Table 33 shows the decimal reduction times (D-values) for the
six most heat stable enzyme preparations.  It was found that
enzyme from Penicillium janthinellum was the most stable among
those studied.  Subsequent investigations of the effect of pH
and salt concentration were done to determine the thermal
stability under conditions which might occur in spent brines.
Figure 5 shows the effect of pH on the D-value of the pectinase
at 75°C (167°F) and 78°C  (172.4°F).  The pH of maximum
stability is 3.7 at both temperatures.  There is a large
increase in stability with only a small change in temperature.
This points up the importance of good temperature control during
brine treatment.  A small drop in temperature could result in
inadequate brine treatment.

     The effect of salt concentration on enzyme stability is
shown in Table 34.  The enzyme is more stable at higher salt
concentrations, but the magnitude of the effect is small.  The
recommended process has a sufficient safety margin included so
that adjustment of the process for the salt concentration of
the spent brine will not be required.

     The thermal stability of P. janthinellum pectinase in
commercial spent brines was less than that an the simulated
brines.  D-values obtained in commercial brines at 12.6% NaCl
and 75°C (167°F) at pH 3.5 and 3.7 were 14 sec and 18 sec,
respectively.  D-values in simulated brines with 12% NaCl, 0.6%
lactic acid and 0.1% Ca++ ion were 29 sec and 33 sec at the
same temperature and pH.

                                58

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62

-------
TABLE 32.  SOURCES AND IDENTITY OF FUNGI USED FOR
           DETERMINATION OF THERMAL STABILITY OF
           PECTINASES IN BRINES
     Organism
 Source*
 Alternaria tenuis

 Cladosporium cladosporioides

 Fusarium oxysporum

 Fusarium roseum

 Fusarium solani

 Penicillium janthinejLlum

 Penicillium oxalicum

 Trichoderma viride
NRRL 2169

  MSU

NRRL 1943

  MSU

NRRL 3078

NRRL 2016

NRRL 790

  MSU
 *NRRL - USDA Northern Regional Research Laboratory,
         Peoria, IL
  MSU - Michigan State University, Department of
        Botany and Plant Pathology.
  TABLE 33.  D-VALUES OF PECTINASES WITH THE GREATEST THERMAL
             STABILITY IN PRELIMINARY EXPERIMENTS.  D-VALUES
             WERE OBTAINED AT pH 3.3 IN 12% NaCl, 0.6%
             LACTIC ACID AND 0.1% Ca++ ION
Organism
Penicillium janthinellum
Penicillium oxalicum
Fusarium solani
Fusarium oxysporum
Alternaria tenuis
Trichoderma viride
Temperature
(°C)
78
75
70
70
70
65
65
D-value
(sec)
4
18
18
5
4
25
10
                              63

-------
       30
   to
   o
   a
   LU
   CO
   LU
   1
20
       10
        0
Figure 5.
                                    75'C
        3,0      3,5
                                          4,5
                            pH
    Effect of pH on heat inactivation of Penicillium
    •janthinellum pectinase.  Experiments were done
    in brine containing 12% NaCl, 0.6% lactic acid
    and 0.1% Ca++ ion.
                             64

-------
 TABLE 34.  EFFECT OF NaCl CONCENTRATION ON THE HEAT
            INACTIVATION OF PECTINASE FROM PENICILLIUM
            JANTHINELLUM AT 75°C
    NaCl, %                  D-values at 75°C  (sec)

8
12
16
pH 3.5
24
29
30
pH 3,
31
33
37
7



TABLE 35.  INACTIVATION OF PECTINASE FROM PENICILLIUM
           JANTHINELLUM AT HIGH pH IN SPENT BRINE.
           TEMPERATURE 22°C (71.6°F).  SALT CONCENTRATION
           8.0%
pH Of
incubation
10.6
11.0
11.2
11.3
11.6
Time for 90%
inactivation
(hr)
50% inact. in 70 hr
34
5.0
6.5
2.5
Time for 99%
inactivation
(hr)

*
37
27
13 (est.)
 *99% inactivation of the enzyme was not reached in 48 hr,
                             65

-------
     At 78°C (172.4°F), the maximum D-value observed under any
of the conditions investigated was 8 sec.  Currently, the
minimum recommended process for brine treatment is 79.4°C
(175°F) for 30 sec in a plate heat exchanger.  This will result
in at least a 30 sec/8 sec = 3.7 D process.  Therefore, at
79.4°C for 30 sec, the extent of inactivation will be 103.7 or
about 5,000-fold.  Since the suggested heating temperature is
1.4°C higher than the highest temperature investigated in
these experiments and the pectinase was significantly less
stable in commercial spent brines, the extent of enzyme
inactivation in commercial brine would be expected to be
considerably greater than 5,000-fold.

     Pectinases are known which are considerably more resistant
to thermal inactivation than 1?. janthinellum enzyme.  A
commercial enzyme from Aspergillus niger was found to have a
D-value of 34 sec at 80°C (176°F) in pH 3.6 simulated spent
brine  (8).  Archer and Fielding  (32) reported a polygalacturon-
ase from Sclerotinia fructigena, which caused disintegration
of fruits after canning, that had a D-value near 3.5 hr when
heated at 90°C in pH 7.0, 0.02 M phosphate buffer.  Therefore,
the possibility exists that a highly stable softening enzyme
could occur in fermentation brines.  However, heating at 79.4°C
(175°F) for 30 sec will be sufficient to protect against
softening from pectinases which are likely to be present on
cucumber fruits and flowers.

     Experiments were also done to determined the stability of
P_. janthinellum pectinase at the high pH levels encountered
during base treatment.  It was found that the enzyme could not
be adequately inactivated if the temperature was held at 10°C
(50°F).  In northern locations, such as Michigan, chemical
treatment should not be done until the brines have warmed up.
In all cases pectinase inactivation at high pH did not show
first order denaturation kinetics.  The rate of inactivation
declined with time.  As a result, meaningful D-values could not
be calculated.  Table 35 shows the effect of different pH levels
at 22°C  (72°F) on the time required for 90% and 99% enzyme
inactivation.  Based upon these results, it is recommended that
brine be held at 72°F or higher during chemical treatment.  The
pH should be raised to 11.2 or higher and held for at least
37 hr at that pH before acidifying the brine.

     Enzymatic softening in Michigan is not common.  Analysis of
spent brines, according to the procedure developed by Bell and
Etchells  (11), after cucumbers were removed from fermentation
tanks, showed no measurable pectinase activity.  As a result, it
was not possible to evaluate the effect of heat and chemical
treatments on the inactivation of pectinase as they occurred in
commercial brines.
                               66

-------
Lyssinoalanine in Base Treated Brine

     Since it was known that base treatment of protein could
lead to formation of lysinoalanine (26) and it had been
reported that free lysinoalanine was toxic to rats  (27), an
experiment was done to determine whether lysinoalanine was
formed during base treatment of spent brine.  A sample of first
cycle brine was adjusted to pH 11.4 and held at room tempera-
ture for 8 days.  The clear supernatant was removed and adjusted
to pH 4.6 with acetic acid.  This treatment was a severe
treatment relative to that required for pectinase inactivation.
Lysinoalanine was found in the spent brine before treatment at
a level of 15 yg/1.  After treatment, 12 yg/1 were detected.
The results indicated that significant amounts of lysinoalanine
were not formed as a consequence of base treatment.  Lysino-
alanine has been found in a number of foods and food proteins
(33, 34).  The level in pickle brine would appear to be quite
low compared to the levels in processed foods which contain
significant levels of protein.
                               67

-------
                           REFERENCES

1.  Anonymous.   Annual Report of Intake  and Inventory  as  of
    October 1,  1975.   Pickle Packers International,  Inc.,
    St.  Charles, Illinois  60174,  1977.

2.  Kirk, D.  G.  Management of Wastewater Problems  in  the Pickle
    Industry.  First  International Congress on Engineering and
    Food, Digest of Papers.  American  Society of Agricultural
    Engineers,  St.  Joseph,  Michigan 49085, 1976, pp.  151.

3.  Etchells, J. L. and Moore, W.  R.,  Jr.   Factors  Influencing
    the  Brining of  Pickling Cucumbers—Questions and Answers.
    Pickle Pak Sci. 1:1, 1971.

4.  Durkee, E.  L.,  Lowe, E., Baker, K. E., and Burgess,  J.  W.
    Field Tests of  Salt Recovery System  for Spent Brine.
    J. Food Sci. 38:507, 1973.

5.  Durkee, E.  L.,  Lowe, E. and Toocheck,  E.  A.  Use of  Recycled
    Salt in Fermentation of Cucumber Salt Stock. J. Food Sci.
    39:1032,  1974.

6.  Geisman,  J. R.  and Henne, R. E. Recycling Food Brine
    Eliminates Pollution.  Food Engr.  45 (1):119,  1973.

7.  Geisman,  J. R.  and Henne, R. E. Recycling Brine from
    Pickling.  Ohio Report 58:76,  1973.

8.  Palnitkar,  M. P.  and McFeeters, R. F.   Recycling Spent
    Brines in Cucumber Fermentations.  J.  Food Sci.  40:1311,
    1975.

9.  Cranfield,  D.  Cucumber Brining and  Salt Recovery.  Pickle
    Packers International Seminar on Pickle Processing Wastes,
    1973.

10. Etchells, J. L.,  Bell,  T. A.,  Fleming, H. P.,  Kelling, R. E.,
    and Thompson, R.  L.  Suggested Procedure for the Controlled
    Fermentation of Commercially Brined  Pickling Cucumbers—the
    Use of Starter Cultures and Reduction of Carbon Dioxide
    Accumulation.  Pickle Pak Sci. 3:4,  1973.
                               68

-------
11. Bell, T. A., Etchells, J.  L.  and Jones,  I.  D.   A Method for
    Testing Cucumber Salt Stock Brine for Softening Activity.
    U.S. Dept.  of Agric., ARS  72-5,  1955, pp.  1-15.

12. Continental Can Company, Inc.  Reference Guidebook for
    Sensory Testing, 3rd ed.,  Chicago, Illinois,  1966.

13. Standard Methods for the Examination of Water and Wastewater,
    14th ed., sec. 507, American Public Health Association,
    New York, 1975, pp. 543-550.

14. Standard Methods for the Examination of Water and Wastewater,
    14th ed., sec. 508, American Public Health Association,
    New York, 1975, pp. 515-554.

15. Standard Methods for the Examination of Water and Wastewater,
    14th ed., sec. 208D, American Public Health Association,
    New York, 1975, p.  94.

16. Miller, G.  L.  Use  of Dinitrosalicylic Acid for Determin-
    ation of Reducing Sugars.   Anal. Chem. 31:426,  1959.

17. Lillevik, H. A.  The Determination of Total Organic Nitrogen.
    Methods in Food Analysis,  2nd ed., Joslyn,  M.  A., ed.,
    Academic Press, New York,  1970,  pp. 601-616.

18. Sprague, S. and Slavin, W.  Determination of  Very Small
    Amounts of Copper and Lead in KC1 by Organic  Extraction and
    Atomic Absorption Spectrophotometry.  Atomic  Absorption
    Newsletter 3:37, 1964.

19. Thomas, B., Roughan, J. A. and Watters,  E.  Lead and
    Cadmium Content of  Some Vegetable Foodstuffs.   J. Sci. Food
    Agr. 23:1493, 1972.

20. Hatch, W. R. and Ott, W. L.   Determination of Sub-Microgram
    Quantities of Mercury by Atomic Absorption Spectrophoto-
    metry.  Anal. Chem. 40:2085,  1968.

21. Chaplin, M. H. and  Dixor^,  A.  R.   A Method for Analysis of
    Plant Tissues by Direct Reading Spark Emission Spectroscopy.
    Appl. Spectroscopy  28:5, 1974.

22. Luke, M. A., Froberg, J. E.  and Masumoto,  H.  T.  Extraction
    and Cleanup of Organochlorine, Organophosphate, Organoni-
    trogen, and Hydrocarbon Pesticides in Produce for Determin-
    ation by Gas-Liquid Chromatography.  JAOAC 58:1020, 1975.
                               69

-------
ic
23. Etchells,  J.  L.,  Bell,  T.  A.,  Monroe,  R.  J. ,  Masley,  P.  M. ,
    and Demain, A.  L.   Populations and Softening  Enzyme Activity
    of Filamentous  Fungi on Flowers,  Ovaries,  and Fruit of
    Pickling Cucumbers.  Appl.  Microbiol.  6:427,  1958.

24. White, W.  L.  and  Downing,  M.  H.   Coccospora agricola
    Goddard, its  Specific Status ,  Relationships and Cellulolyt
    Activity.   Mycologia 43:656,  1951.

25. Chavana, S.  and McFeeters,  R.  F.   Thermal Inactivation  of
    Fungal Pectinases in Cucumber Brines.   Lebensm.-Wiss . U.
    Technol. 10:290,  1977.

26. DeGroot, A.  P.  and Slump,  P.   Effects  of  Severe Alkali
    Treatment of  Proteins on Amino Acid Composition and
    Nutritive Value.   J. Nutrition 98:45,  1969.

27. Woodward,  J.  C. and Short,  D.  D.   Toxicity of Alkali-Treated
    Soy Protein in  Rats.  J. Nutrition 103:569, 1973.

28. Costilow,  R.  N.,  Bedford,  C.  L. ,  Mingus ,  D. and Black,  D.
    Purging of Natural Salt-Stock Pickle Fermentations  to
    Reduce Bloater  Damage.   J.  Food Sci. 42:234,  1977.

29. Horngren,  C.  T.  Cost Accounting — a Managerial Emphasis.
    4th ed. , Prentice-Hall, Englewood Cliffs, New Jersey, 1977.

30. United States Environmental Protection Agency.  Development
    Document for Interim Final and Proposed Effluent Limitations
    Guidelines and  New Source  Performance  Standards for  the
    Fruits, Vegetables and Specialties Segment of the Canned
    and Preserved Fruits and Vegetables Point Source Category.
    EPA 440/1-75/046, 1975.

31. Anonymous.  Canned and Preserved Fruits and Vegetables  Point
    Source Category.   Federal  Register 41  (75):16272, 1976.

32. Archer, S. A. and Fielding, A. H.  Thermostable Polygalac-
    turonase Secreted by Sclerotinia fructigena.   J. Food Sci.
    40:423, 1975.

33. Sternberg, M. ,  Kim, C.  Y.  and Schwende, F. J.  Lysinoalanine:
    Presences in Foods and Food Ingredients.   Science 190:992,
    1975.

34. Chu, N. T., Pellett, P. L.  and Nawar,  W.  W. Effect  of
    Alkali Treatment  on the Formation of Lysinoalanine  in Corn.
    J. Ag. Food Chem. 24:1084,  1976.

35. Pesticide Analytical Manual,  vol. I, Food and Drug  Admin-
    istration, Washington,  DC,  Rev.  March, 1977,  sec. 211. 14d,
    1968.

                               70

-------
TABLE A-l.
             APPENDIX

COMPOSITION OF FIRST CYCLE SPENT BRINE BEFORE
     HEAT TREATMENT, 1975 DATA
Tank number
Parameter
Acid % lactic
Salt %
pH
Sugar mg/1
SS mg/1
BOD mg/1
COD mg/1
Kjeldahl N mg/1
Cd mg/1
Hg mg/1
P mg/1
Ca mg/1
Mg mg/1
Mn mg/1
Fe mg/1
Cu mg/1
B mg/1
Zn mg/1
Al mg/1
Cr mg/1
K7
.319
12.10
3.52
325
238
7550
12745
517
.031
0
119
1010
130
3.6
4.9
—
1.4
6.8
6.3
.012
Kll
.577
12.96
3.52
585
240
14000
15900
738
—
0
154
980
150
3.0
7.5
1.8
1.8
5.4
5.3
.001
K41
.360
10.92
3.57
306
200
8250
11235
499
.013
0
102
910
130
3.3
4.9
1.7
1.3
5.1
5.3
0
L8
.420
14.02
3.68
404
300
10500
14195
597
0
0
131
980
130
2.4
7.5
1.7
1.4
4.7
4.7
0
L39
.480
14.10
3.58
391
300
9800
16500
712
.016
0
142
1090
160
1.9
6.9
1.5
1.3
3.8
5.3
0
L40
.412
13.64
3.57
427
185
10450
13855
612
.005
0
114
930
140
1.1
3.7
0.9
1.0
2.8
4.2
0
L43
.501
14.28
3.68
403
320
11700
15025
530
.027
0
148
1170
150
3.6
8.1
2.0
1.6
5.6
6.3
.010
                            71

-------
TABLE A-2.  COMPOSITION OF FIRST CYCLE SPENT BRINE
          AFTER HEAT TREATMENT, 1975 DATA
Tank number
Parame'ter
Acid % lactic
Salt %
pH
Sugar mg/1
SS mg/1
BOD mg/1
COD mg/1
Kjeldahl N mg/1
Cd mg/1
Hg mg/1
P mg/1
Ca mg/1
Mg mg/1
Mn mg/1
Fe mg/1
Cu mg/1
B mg/1
Zn mg/1
Al mg/1
Cr mg/1
K7
.087
12.50
4.72
228
138
10375
13080
559
.008
0
119
1060
130
3.6
8.7
1.8
1.7
5.8
17
0
K41
.111
11.00
4.73
273
273
8700
11560
495
0
0
102
910
120
3.3
7.5
1.8
1.3
5.1
9.0
.003
K41
.208
11.16
4.39
276
195
9250
11815
500
.016
0
91
810
120
1.1
5.8
1.2
1.0
2.6
7.4
.003
L8
.074
12.20
5.02
123
100
7825
10825
395
0
0
80
960
100
3.3
4.9
2.1
1.0
5.1
13.8
0
L39
.229
14.38
4.45
271
336
12000
15460
—
.179
0
148
1060
150
3.3
9.0
1.6
1.7
4.4
15.9
.010
L40
.331
13.96
4.76
463
185
10300
13910
585
.027
0
131
1040
150
1.9
6.4
1.2
1.5
3.3
11.6
0
                        72

-------
TABLE A-3.  COMPOSITION OF FIRST CYCLE SPENT BRINE
        BEFORE CHEMICAL TREATMENT, 1975 DATA
Tank number
Parameter
Acid % lactic
Salt %
pH
Sugar mg/1
SS mg/1
BOD mg/1
COD mg/1
Kjeldahl N mg/1
Cd mg/1
Hg mg/1
P mg/1
Ca mg/1
Mg mg/1
Mn mg/1
Fe mg/1
Cu mg/1
B mg/1
Zn mg/1
Al mg/1
Cr mg/1
K10
.398
13.29
3.48
408
125
9850
13620
486
.162
0
114
930
120
2.7
1.6
2.0
1.2
4.2
5.3
.012
L6
.334
13.39
3.46
305
300
7850
10545
—
—
0
102
1040
130
3.3
7.8
1.9
1.5
4.9
5.3
0
L9
.410
14.11
3.66
490
310
10200
13820
600
.013
0
125
1060
130
2.7
6.9
1.6
1.3
4.4
4.7
0
L42
.563
14.66
3.58
484
267
12300
16135
637
.179
0
114
1010
130
3.8
8.1
2.2
1.3
5.6
7.9
.003
M8
.650
10.86
3.50
280
155
12560
16085
536
.019
0
125
1060
140
4.4
8.4
2.3
1.6
5.8
7.4
0
                         73

-------
TABLE A-4.  COMPOSITION OF  FIRST  CYCLE SPENT BRINE
         AFTER CHEMICAL TREATMENT,  1975 DATA
Tank number
Parameter
Acid % lactic
Salt %
PH
Sugar mg/1
SS mg/1
BOD mg/1
COD mg/1
Kjeldahl N mg/1
Cd mg/1
Hg mg/1
P mg/1
Ca mg/1
Mg mg/1
Mn mg/1
Fe mg/1
Cu mg/1
B mg/1
Zn mg/1
Al mg/1
Cr mg/1
L41
.415
10.38
4.60
151
35
12300
15665
457
.023
0
11
570
10
0.5
1.7
0.7
1.0
1.7
6.4
0
L42
.289
10.60
4.55
170
55
15750
18265
516
.001
0
28
470
20
0.5
2.9
0.4
0.7
1.3
7.9
0
L47
.208
14.07
5.10
349
22
16625
21620
616
.009
0
34
810
10
2.4
2.0
1.4
1.6
3.8
6.3
0
M45
.058
13.50
5.25
194
110
8750
12700
432
.098
0
17
810
10
2.2
1.4
1.6
1.1
3.8
13.8
.003
L7
.192
14.37
5.00
242
53
16650
17795
594
.159
0
40
910
10
4.1
4.6
2.2
1.8
5.4
23.5
0
                          74

-------
TABLE A-5.  COMPOSITION OF  2ND CYCLE SPENT
            AND AFTER HEAT  TREATMENT,  1976
BRINE BEFORE
DATA
Tank number
Parameter
Acid % lactic
Salt %
pH
Sugar mg/1
SS mg/1
BOD mg/1
COD mg/1
Kjeldahl N mg/1
Cd mg/1
Hg mg/1
P mg/1
Ca mg/1
Mg mg/1
Mn mg/1
Fe mg/1
Cu mg/1
B mg/1
Zn mg/1
Al mg/1
Before
L46
.57
10.7
3.52
230
157
10700
17592
540
.03
.003
119
910
170
0.5
3.4
0.8
1.1
2.8
3.2
M4
.66
11.5
3.55
276
160
13400
19768
641
.03
.003
165
1090
180
1.1
14.7
1.1
1.3
4.0
9.0
treatment
M8
.63
11.9
3.52
319
252
13500
19504
614
.01
.003
159
1110
180
2.4
11.7
2.0
1.5
6.6
10.0
M10
.61
11.6
3.50
227
622
12400
17576
540
.03
.003
114
1060
160
1.6
7.5
1.7
1.5
5.6
11.1
M49
.20
12.3
4.58
232
220
13200
18724
631
.02
.003
159
1220
190
1.6
9.9
1.8
1.5
5.4
20.2
After treatment
M4
.22
11.8
4.49
205
100
12363
18720
571
.02
.004
108
1220
180
1.6
5.8
1.5
1.6
5.4
7.4
M3
.30
11.8
4.33
262
194
14000
20128
669
.03
.003
137
1140
180
2.4
23.9
1.6
1.7
7.3
14.3
L44
.12
11.5
5.10
189
313
16600
19616
613
.025
.003
108
1010
180
1.1
5.2
1.4
1.5
4.4
6.3
                       75

-------
TABLE A-6.  COMPOSITION OF 2ND CYCLE  SPENT  BRINE BEFORE
       AND AFTER CHEMICAL TREATMENT,  1976 DATA
Parameter
Acid % lactic
Salt %
PH
Sugar mg/1
SS mg/1
BOD mg/1
COD mg/1
Kjeldahl N mg/1
Cd mg/1
Hg mg/1
P mg/1
Ca mg/1
Mg mg/1
Mn mg/1
Fe mg/1
Cu mg/1
B mg/1
Zn mg/1
Al mg/1


Before
L47
.65
9.6
3.60
292
179
13300
18552
453
.025
.003
85
760
100
0.5
20.5
0.8
0.9
3.1
4.2
M6
.57
11.3
3.82
259
167
12700
19544
608
.02
.003
97
1010
160
1.1
15.9
0.8
1.5
4.4
4.7

Tank
treatment
M43
.57
12.3
3.58
228
117
13600
18032
590
.03
.003
102
1010
140
1.1
17.7
1.7
1.4
4.7
14.8
M48
.74
11.2
3.67
253
170
18400
21704
838
.02
—
114
1060
160
1.1
15.9
0.8
1.5
4.4
4.7
number
After treatment
L47
.04
10.0
6.65
87
104
18817
18667
465
.01
.003
11
640
10
0.5
1.1
1.7
1.3
3.3
32.8
M5
.31
10.4
4.80
90
208
14800
23152
543
.05
.003
11
590
10
0.3
2.3
1.3
1.5
2.6
21.3
Mil
.24
11.6
4.90
104
14
15921
22368
489
.03
.003
11
810
0
1.1
2.6
1.4
1.5
4.2
36.1
M5
.43
11.4
4.80
178
22
19840
26952
531
.03
.003
11
570
0
0.5
13.5
2.0
1.7
4.4
34.4
                           76

-------
TABLE A-7.  ANALYSIS OF  1ST  CYCLE  CONTROL SPENT
       BRINES AFTER REMOVAL  OF  SALT STOCK
Tank no.
-1975-
L36
M2
M51
Ml
M52
M9
M42
M35
M33
L48
-1976-
M7
M50
M47
M46
M6
M43
M9
M8
Acid
%
.670
.742
.640
.657
.669
.761
.649
.718
.716
.498
.568
.514
.782
.557
.579
.520
.415
.629
Salt
%
8.6
9.7
10.5
12.1
8.5
10.2
8.4
9.6
9.7
8.7
9.3
8.1
10.2
7.7
9.4
7.6
6.7
9.5
PH
3.30
3.40
3.35
3.32
3.39
3.29
3.38
3.38
3.37
3.50
3.52
3.62
3.57
3.60
3.50
3.60
3.50
3.49
Reducing
sugar
mg/1
378
575
571
125
193
493
481
505
168
176
172
134
258
208
203
166
140
210
BOD
mg/1
11480
15900
15010
13800
13657
17000
11375
17200
21400
10000
14000
10800
18800
14558
12000
11612
6600
12000
COD
mg/1
15736
20296
15864
17520
16525
18880
18500
19260
18575
14825
16990
14309
23550
18116
17184
15131
12210
19066
Suspended
solids
mg/1
227
540
463
488
160
311
—
388
288
213
347
820
480
730
961
420
127
535
                        77

-------
TABLE A-8.  ANALYSIS OF  2ND AND  3RD CYCLE HEAT TREATMENT
        SPENT BRINES AFTER REMOVAL OF SALT STOCK
Tank no.
-1975-
(2nd cycle)
L37
M3
M49
M47
M5
M7
M10
M41
Mil
M8
L46
-1976-
(3rd cycle)
Ml
M52
M3
M44
M49
M42
M4
L48
Acid
%
.628
.892
.564
.732
.623
.534
.749
.852
.645
.702
.551

.751
.767
.833
.738
.887
.750
.824
.371
Salt
%
10.
8.
12.
10.
8.
9.
12.
12.
11.
9.
7.

7.
12.
9.
9.
9.
11.
10.
6.
2
8
2
2
7
0
0
2
3
3
7

3
6
7
9
6
5
1
5
pH
3.
3.
3.
3.
3.
3.
3.
3.
3.
3.
3.

3.
3.
3.
3.
3.
3.
3.
3.
43
50
50
60
67
51
52
25
45
48
55

64
62
62
64
80
60
55
83
Reducing BOD
sugar mg/1
mg/1
467
479
527
76
420
370
499
361
531
431
402

179
181
192
186
172
186
191
119
13480
20200
13600
19000
6820
9320
21400
10200
18700
12720
12400

18763
20400
20400
18400
23800
21400
19600
11000
COD
mg/1
16584
25600
16730
21695
17824
15584
23535
11390
20170
19112
17180

22935
25555
27184
24506
27776
25396
24850
12846
Suspended
solids
mg/1
76
967
1025
675
765
124
500
330
750
207
660

175
771
617
732
271
230
185
53
                            78

-------
TABLE A-9.  ANALYSIS  OF 2ND AND 3RD CYCLE CHEMICALLY  TREATED
          SPENT BRINES  AFTER REMOVAL OF SALT STOCK
Tank no.
-1975-
(2nd cycle)
L44
M4
M50
M48
M6
M43
M40
M39
M45
L47
-1976-
(3rd cycle)
M2
M51
M48
M45
M5
M41
Mil
L47
Acid
%
.852
.787
.642
.890
.755
.697
.723
.636
.594
.750


.937
.507
.978
.538
1.14
.505
.770
.806
Salt
%
10
8
10
10
9
8
9
10
9
7


7
9
10
7
10
7
8
8
.6
.8
.0
.8
.6
.4
.6
.1
.1
.3


.7
.1
.6
.6
.2
.2
.6
.8
3
3
3
3
3
3
3
3
3
3


3
3
3
3
3
3
3
3
pH
.50
.62
.65
.65
.63
.58
.38
.60
.70
.59


.78
.73
.78
.70
.65
.72
.41
.58
Reducing BOD
sugar mg/1
mg/1
512
169
380
431
410
442
69
71
342
420


258
162
350
208
328
184
311
274
17600
17000
15800
20300
14520
13250
17300
17000
14800
15600


21000
13000
25400
13275
27200
13600
21000
22600
COD
mg/1
23288
23490
18910
24815
22104
18732
18945
18055
16604
19670


26082
15667
32323
16351
30784
15028
23508
25068
Suspended
solids
mg/1
172
—
457
417
136
—
275
380
144
139


367
1610
880
660
820
387
90
80
                              79

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-------
TABLE A-30.  ACID PRODUCTION DURING  1ST  CYCLE  FERMENTATION
    OF CUCUMBERS USING FRESH CONTROL BRINE—1976  DATA.
    RESULTS ARE EXPRESSED AS PERCENT TITRATABLE ACIDITY
                 CALCULATED AS LACTIC ACID
Fermentation
time (days)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
Tank number
M50
	
.01
.04
	
.13
.17
.24
.29
.33
.42
.45
.50
.51
.50
.60
.55
.65
.68
.70
.66
.69
.63
M7
	
.05
.05
	
.16
.23
.30
.39
.39
.49
.50
.54
.58
.63
.69
.68
.70
.75
.74
.78
.83
.75
M47
	
.02
.02
.04
.06
.21
.25
.29
.32
.36
.38
.50
.55
.60
.62
.64
.76
.68
.71
.75
.85
M46
.02
.05
.03
.05
.10
.18
.29
.36
.42
.46
.52
.54
.56
.67
	
.72
.65
	
.66
.63
M6
.02
.06
.16
.24
.29
.34
.43
.54
.64
.76
.77
	
.83
.78
.83
	
	
.82
.84
.92
.93
.91
M43
	
.02
.04
.05
.22
.07
.16
.21
.25
	
.40
.40
.44
.27
.34

.61
.58
.61
.65
.65
	
M9
	
.05
.07
.10
	
.37
.37
.47
.56
.62
	
	
.51
.68
.79
.78
.80
	
	
.87
.84
M8
	
.04
.14
.23
.31
.34
	
	
.47
.61
.64
.66
.66
	
	
	
.77
.75
.70
	
	
                            100

-------
TABLE A-31.  ACID PRODUCTION DURING 1ST CYCLE FERMENTATION
    OF CUCUMBERS USING  FRESH CONTROL BRINE—1975 DATA.
    RESULTS ARE EXPRESSED  AS PERCENT TITRATABLE ACIDITY
                 CALCULATED AS LACTIC ACID
Fermentation
time (days)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
L36

.03
.15
.27
	
.48
.57
.57
.63
.63
.60
	
.81
.84
.90
.87
.87
.90
.96
.93
1.02
.96
M2

.03
.09
.15
.24
.27
.45
.48
.54
.60
.66
.78
.60
.66
.72
.78
.63
.81
.84
.78
.69
.78
M51

.06
.12
.18
.54
.45
.51
.54
.60
.66
.63
.62
.75
.75
.75
.78
.78
.66
.78
.81
.69
.72
Tank number
Ml

.03
.06
.09
.18
.21
.36
.46
.33
.42
.51
.57
.57
.63
.69
.75
.69
.72
.63
.69
.75
.60
M52

.03
.06
.21
.36
.57
.60
.54
.60
.63
.63
.63
.69
.78
.78
.81
.78
.78
.81
.87
.87
.81
M9

.06
.18
.57
.54
.57
.57
.60
.66
.63
.69
.78
.78
.81
.78
.81
.81
.90
.87
.81
.93
.84
M42

.03
.09
.24
.30
.45
.54
.42
.39
.42
.57
.54
.63
.63
.63
.54
.78
.75
.75
.78
.69
.63
M35

.03
.03
.24
.33
.36
.45
.57
.48
.57
.63
.57
.60
.63
.66
.63
	
.69
.72
.72
M33

.03
.18
.27
.12
.18
.24
.33
.42
.48
.54
.60
.63
.63
.69

.72
.78
.78
	
L48

.03
.06
.09
.09
.12
.21
.36
	
.54
.63
.66
	
	
	
.81
.72
.69
.69
	
                             101

-------
  TABLE A-32.   ACID PRODUCTION  DURING 2ND  CYCLE FERMENTATION
  OF CUCUMBERS USING 1ST CYCLE  HEAT TREATED BRINE—1975  DATA.
     RESULTS ARE EXPRESSED AS PERCENT TITRATABLE ACIDITY
                   CALCULATED AS LACTIC ACID
Fermentation 	Tank number	
time (days)   L37  M3  M49  M4?   M5  M?  M1Q  M41  Mll Mg   L46

     1
     2       .30 .12  .24  .21  .18 .18  .06  .09  .06 .03   .06
     3       .18 .18  .21  .24  .21 .15  .12  .12  .06 .30   .09
     4       .33 .21  .45  .33  .36 .39  .42  .24  .27 .15   .12
     5       .54 .36  .45  .57  .48 .60  .54  .39  .42 .30   .21
     6       .60 .81  .57  .45  .57 .63  .63  .42  .48 	   .30
     7       .60 .51  .66  .57  .66 .69  .69  .48  .57 .54   .42
     8       .72 .66  .75  .63  .69 .75  .75  .54  .57 .57   .54
     9       .75 .75  .78  .60  .75 .81  .72  .57  .63 .54   	
    10       .75 .78  .90  .48  .84 .75  .72  	  	.60   	
    11       	.81  .87  	.81 .78  .90  .63  .72 .81   .63
    12       .81 .87  .93  .78  .81 .87  .84  .66  .72 .69   .72
    13       .84 .87  .90  .78  .87 .72  .84  .54  .78 .75   .75
    14       .93 .96  .96  .81  .84 .72  .84  .60  .63 —   —
    15       .87 .96 1.02  .81  .84 .75  .90  .54  .81 	   	
    16       .90 .99 1.02  .87  .84 .75  .90  .57  .81 .78   	
    17       .96 .99  .99  .81  .87 .69  .99  .63  .78 .75   	
    18       .931.02 1.05  .84  .87 .75  .96  —  — .84   —
    19       .901.02  .96  .72  .96 .78  .90  —  	   .81
    20      1.021.05  .87  .84  .87 .72  .99  .33  .81	   .87
    21       .991.11  .93  .90  .84 .84  .93  .39  .87	   .84
    22      1.021.08  .87  .78  .99 .75  .96  .48  .75 —   —
                              102

-------
TABLE A-33.  ACID  PRODUCTION DURING 2ND CYCLE FERMENTATION
  OF CUCUMBERS USING 1ST CYCLE CHEMICAL TREATED BRINE—-
1975 DATA.  RESULTS  ARE  EXPRESSED AS PERCENT TITRATABLE
          ACIDITY  CALCULATED AS LACTIC ACID
Fermentat
time (day
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
99
ion
s)
L44

.24
.33
.51
	
.69
.75
.71
.87
.81
.87
	
.96
1.05
1.23
.99
.99
1.05
1.05
1.02
1.08
1 OR
Tank number
M4

.30
.39
.42
.51
.60
.60
.66
.75
.78
.81
.87
.84
.90
.93
.96
1.02
.99
1.02
1.05
.96
i n=;
M50

.39
.12
.24
.30
.54
.69
.72
.75
.81
.75
.84
.90
.90
.99
1.05
.96
1.02
1.02
.96
.99
QQ
M48

.15
.12
.21
.42
.36
.66
.75
.81
.90
.96
.99
1.02
.99
1.05
1.08
1.11
1.14
1.05
1.08
1.17
1 rm
M6

.09
.18
.60
.63
.78
.87
.87
.93
.99
.96
.93
.96
1.05
.96
.99
.96
.96
.99
1.08
.99
m
M43

.12
.24
.54
.75
.60
.69
.72
.69
.78
.90
.81
.93
.93
.93
.90
.99
.99
.87
.84
.96
QT
M40

.06
.06
.24
.33
.27
.39
.39
.42
.51
.54
.60
.63
.66
.69
.66
___
.60
.69
•7=;
M39

.06
.06
.33
.42
.48
.33
.33
.54
.63
.69
.57
.63
.75
.78
.78
___
.75
.84
RA
M45

.09
.09
.09
.18
	
.36
.57
.54
.60
.66
.66
.54

.75
.81
.84
	
	
L47

.18
.21
.33
.48
.54
.60
.69
.84
.90
.93

__—
.96
.99
1.05
.99
                             103

-------
TABLE A-34.  ACID PRODUCTION DURING 3RD CYCLE FERMENTATION
OF CUCUMBERS USING 2ND CYCLE HEAT TREATED BRINE—1976 DATA.
  RESULTS ARE EXPRESSED AS PERCENT TITRATABLE ACIDITY
                CALCULATED AS LACTIC ACID
Fermentat
time (day
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
ion
S) M52
_ —
.26
.23
.29
.43
.53
.59
.69
.74
.73
.77
.81
.79
.84
.84
.88
.90
.91
Tank number
Ml
.24
.23
	
.49
.61
.61
.64
.67
.73
.72
.74
.77
.78
.84
.85
.83
.85
.83
M3
	
.16
.15
.16
.26
.41
.49
.60
.62
.64
.76
.79
.79
.89
.91
.84
.90
.73
.84
M44
.12
.14
.12
.08
.28
.32
.38
.44
.50
.60
.65
.70
.69
.75
.85
.80
.71
	
M49
.13
.19
.34
.46
.56
.61
.70
.70
.86
.91
.97
.96
.69
.80
_ —
.87
.87
M42
-__
.16
.17
.17
.26
.31
.36
.37
.42
.63
.59
.66
.49
.60
___
.63
.64
M4
___
.13
.13
.15
.41
.49
.49
.59
.62
	
.73
.76
.82
.84
.83
	
	
L48
	
.15
.30
.44
.48
.57
	
	
.84
.87
.95
.92
.97
	
1.01
	
1.03
                            104

-------
TABLE A-35.  ACID PRODUCTION DURING 3RD CYCLE FERMENTATION
  OF CUCUMBERS USING  2ND  CYCLE  CHEMICAL TREATED BRINE —
 1976 DATA.  RESULTS  ARE  EXPRESSED AS PERCENT TITRATABLE
            ACIDITY CALCULATED  AS LACTIC ACID
Fermentation
time (days)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
M51
	
.23
.23
	
.30
.45
.54
.64
.69
.80
.78
.82
.84
.88
.87
.78
.80
.83
.89
M2
.25
.24
	
.41
.54
.61
.75
.79
.85
	
.93
.99
1.01
1.06
1.07
1.08
1.10
1.08
1.06
M48
	
.20
.17
.18
.16
.33
.42
.51
.55
.61
.70
.75
.81
.80
.89
.88
.98
.93
.91
Tank
M45
.15
.09
.14
.07
.23
.36
.49
.58
.67
.70
.71
.61
.73
.80
	
.91
.80
	
.81
number
M5
.12
.19
.38
.56
.66
.73
.79
.80
.91
	
1.04
1.13
1.06
1.10
1.11
	
	
1.11
1.11

M41
.12
.09
.15
.34
.36
.42
.46
.50
	
.75
.78
.78
.62
.69
.67
.77
.90
.82
.86

Mil
	
.12
.13
.16
	
.46
.47
.56
.63
.71
.81
.88
.98
.97
1.03
	
	
	
1.02

L47
	
.21
.35
.48
.63
.63
.82
.79
.91
.88
.91
	
	
	
.88
.82
.91
	
___
                             105

-------
TABLE A-36.  INSTRON PRESSURE TEST ON  2ND CYCLE  SALT STOCK
    CUCUMBERS AFTER DESALTING.  RESULTS ARE EXPRESSED
    AS kg OF FORCE REQUIRED TO PUNCTURE THE FRUIT WITH
               A 5/16" DIAMETER PROBE
Group no. Control
1 6.07
2 8.67
3 4.77
4 8.63
5 8.80
6 9.19
7 7.65
8 8.60
9 8.84
10 	
11 8.09
Heated brine
8.28
7.98
8.75
8.86
7.87
8.67
8.85
8.64
9.24
7.55
7.08
Chemically
treated brine
8.88
8.92
7.35
8.40
7.62
	
7.61
9.04
9.60
7.40
8.48
                             106

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 TABLE A-37.  SAMPLE CALCULATIONS FOR 1 BUSHEL CUCUMBER
                    SALTING (DESALTING*)


        Item                                   Amount

Fermentation                              Ib Salt   Total Ib

   Ib salt in 3.2 gal cover brine at 25°S    1.84     	
   Ib salt required to maintain 25°S         3.46     	

Storage

   Ib salt required to raise salt concen-
      tration to 45°S                        4.60     9.90
Untanking

   Ib salt remaining in 3.8 gal spent
      brine                                  4.09     	

   Ib salt in 1 bushel or 5.4 gal
      pickles                                5.81     9.90

Desalting

   Ib salt in desalted pickles               2.21     	

   Ib salt lost in process water             3.60     	

Summary                                  No recycling Recycling
   total Ib salt to 45°S                     9.90     9.90

   Ib salt in spent brine                    4.09     4.09

   Ib salt in process water                  3.60     3.60

   Ib salt in excess brine (after
fermentation)
Ib salt saved because of recycling
Ib salt to waste treat
0.65
	
7.69
0.65
3.44
4.25
*Basis:  1. Bushel occupies 6 gal volume
         2. 3.2 gal needed to cover 1 bushel
         3. 65 parts cucumbers, 35 parts brine by weight

         4. 10% shrinkage of pickle stock, thus 10%
            excess brine generation

         5. Stock stored at 45°S desalted to 18°S.
                             107

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TABLE A-38.  HEAT INACTIVATION OF FUNGAL PECTINASES  AT pH 3.3
       IN 12% NaCl, 0.6% LACTIC ACID AND 0.1%  Ca++ ION
Organism
Penicillium janthinellum


Penicillium janthinellura



Penicillium oxalicum



Fusarium solani



Fusarium oxysporum



Alternaria tenuis



Trichoderma viride



Heating
temperature
(°C)
78


75



70



70



70



65



65



Heating
time
(sec)
20
24
25
30
40
40
45
30
40
45
50
15
17
18
20
18
20
22
23
20
30
45
60
30
35
40
45
Activity
remaining
(%)
16.3
1.9
1.1
11.2
2.9
2.8
1.6
32.7
9.6
3.2
3.0
17.0
6.0
3.5
1.5
27.0
9.0
2.6
1.5
51.1
20.7
4.6
1.3
22.7
10.6
8.4
5.0
                              108

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TABLE A-39.  EFFECT OF pH ON INACTIVATION  OF  Penicillium
        janthinellum PECTINASE AT  75°C

PH
3.1



3.3



3.5




3.7



4.0


4.5


4.7




Heating
time (sec)
28
30
33
35
25
30
35
40
30
35
37
40
45
35
40
45
50
35
40
45
30
32
35
25
27
30
32
35
Activity
remaining (%)
6.2
4.1
2.7
1.8
19.0
10.7
6.6
3.3
10.3
6.5
5.5
4.4
3.1
10.3
6.9
5.1
3.6
27.9
15.5
8.2
15.5
10.2
4.3
33.3
19.9
7.7
3.2
1.4
                           109

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Table A-40.  EFFECT OF pH ON INACTIVATION OF Penicillium
     janthinellum PECTINASE AT 78°C
        pH           Heating            Activity
                    time (sec)        remaining (%)

       3.3             20                 16.3
                       24                  1.9
                       25                  1.1

       3.5             20                 23.0
                       24                  4.7
                       26                  2.0

       3.7             20                 16.3
                       24                  5.8
                       25                  4.0
                       26                  3.3

       4.0             20                 42.1
                       23                 19.7
                       26                  5.3
                       28                  1.8

       4.5             20                 32.1
                       23                  9.0
                       25                  5.3
                            110

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TABLE A-41.  EFFECT OF NaCl CONCENTRATION  OF  THE  HEAT
 INACTIVATION OF Penicillium  janthinellum  PECTINASE
AT 75°C
NaCl (%)
8



12




16




8



12



16




pH Heating time
(sec)
3.5 30
35
40
45
3.5 30
35
37
40
45
3.5 30
35
40
50
60
3.7 30
35
40
45
3.7 35
40
45
50
3.7 30
40
50
60
70
Activity
remaining (%)
12.7
5.0
3.3
1.7
10.3
6.5
5.5
4.4
3.1
32.3
22.3
15.5
6.9
3.4
10.7
6.8
4.8
3.5
10.3
6.9
5.1
3.6
37.8
19.0
11.4
5.8
3.1
                            111

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TABLE A-42.  HEAT INACTIVATION OF Penicillium janthinellum
      PECTINASE IN 12.6% NaCl COMMERCIAL SPENT BRINE
       Brine pH           Heating time       Activity
                             (sec)        remaining  (%)


          3.5                 30              48.2
                              40               9.4
                              45               4.4
                              50               1.9

          3.7                 30              25.4
                              36              11.8
                              45               3.5
                              50               1.9
                            112

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TABLE A-43.  INACTIVATION OF PECTINASE FROM Penicillium
   janthinellum AT HIGH pH IN SPENT BRINE.  TEMPERATURE
22°C
pH
10.6










11.0










11.2





11.3










(7l.6°F). SALT
Incubation
time (hr)
0
1
2
3
5
8
14
23
34
47
70
0
1
2
4
5
7
10
15
24
36
48
0
3
6
22
30
36
0
1
2
3
4
6
9
15
24
36
48
CONCENTRATION 8.0%
Activity
remaining (%)
94.9
79.4
74.4
83.9
68.0
64.8
62.6
46.8
42.0
49.0
52.0
82.0
65.7
57.7
41.9
40.7
34.3
24.3
15.0
11.5
9.9
8.0
75.8
16.1
8.2
5.8
2.2
ND*
81.6
37.6
16.3
13.1
12.2
19.2
8.0
6.4
1.6
0.3
0.6
                        (continued)

                            113

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             TABLE A-43 Continued
pH
11.6




Incubation
time (hr)
0
1
3.5
6.5
22
Activity
remaining (%)
59
22
6.1
3.4
ND*
*ND = Not detectable.
                        114

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TABLE A-44.  INACTIVATION OF Penicillium  janthinellum
 PECTINASE IN SIMULATED SPENT BRINE AT  10*C  (50°F).
             SALT CONCENTRATION 12%
     pH           Incubation         Activity
                  time  (hr)        remaining  (%)
11.0 0
6
10
21
33
45
57
69
11.7 0
2
9
20
30
40
71.9
69.8
69.8
63.3
61.3
59.7
56.5
51.6
70.1
60.4
65.3
55.5
53.5
53.9
                           115

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                                  TECHNICAL REPORT DATA
                           (Please read Instructions on the reverse before completing)
1  REPORT NO.
  EPA-600/2-78-207
                             2.
                                                          3. RECIPIENT'S ACCESSION NO.
4. TITLE AND SUBTITLE

  REUSE OF FERMENTATION BRINES  IN THE CUCUMBER

  PICKLING INDUSTRY
                                5. REPORT DATE
                                 September 1978  issuing date
                                6. PERFORMING ORGANIZATION CODE
7. AUTHOR(S)
  R. F. McFeeters  and W.  Coon*; M. P. Palnitkar  and
  M. Velting**;  N.  Fehringer***
                                                          8. PERFORMING ORGANIZATION REPORT NO.
9. PERFORMING ORGANIZATION NAME AND ADDRESS
  Pickle Packers International
  108 East Main Street
  P.O. Box 31
                                10. PROGRAM ELEMENT NO.

                                     1BB610
                                11. CONTRACT/GRANT NO.
  St.  Charles ,  IL
60174
                                                                S-803825
12. SPONSORING AGENCY NAME AND ADDRESS
   Industrial Environmental Research Laboratory
   Office of Research and  Development
   U.S. Environmental Protection Agency
   Cincinnati, Ohio 45268
                                13. TYPE OF REPORT AND PERIOD COVERED
                                   Final Report  5/75  - 12/77
                                14. SPONSORING AGENCY CODE

                                     EPA/600/12
15. SUPPLEMENTARY NOTES
  *Department of  Food Science and Human Nutrition,  MI  St.  Univ., East Lansing,MI 48824
 **Vlasic Foods,  Inc., West Bloomfield, MI 48033
***Detroit District  Office. U.S. FDA. Detroit. MI  48826	
16. ABSTRACT
       The project evaluated on a  commerical scale the technological  and economic
  feasibility of recycling spent cucumber fermentation brine.  Two  brine treatment
  procedures, heat treatment and chemical treatment, were used.   The  results showed
  that brine recycling was practical  on a commerical scale.  Either brine treatment
  procedure resulted in salt stock which were equivalent in quality to control
  cucumbers.

       Studies were conducted to determine the adequacy of the brine  treatment
  procedures employed.  The data confirmed that a heat treatment  of 175°F for 30
  sec. was sufficient to assure inactivation of pectinases from molds found to be
  common on cucumber fruits and flowers.  For effective chemical  treatment a brine
  temperature of 72°F or higher was required.  In addition, the pH  had to be main-
  tained at 11.0 or higher for at  least 36 hr to assure 99% inactivation of pectin-
  ases from the molds which were investigated.

       An economic evaluation of the  recycling procedures showed  a  small net savings
  for  the heat treatment procedure and  a small net cost for chemical  treatment.
  Selection of the process for a particular plant will depend upon  the local conditions,
17.
                               KEY WORDS AND DOCUMENT ANALYSIS
                  DESCRIPTORS
                                             b.IDENTIFIERS/OPEN ENDED TERMS
                                              c. COS AT I Field/Group
  Food  Processing
  Vegetables
  Brines
  Circulation
  Economic Analysis
  Treatment
                       Cucumber Brine
                       Recycle
                       Commercial Scale
  68 D
18. DISTRIBUTION STATEMENT

  Release to Public
                   19. SECURITY CLASS (ThisReport)
                      Unclassified
21. NO. OF PAGES
      130
                                              20. SECURITY CLASS (Thispage)
                                                 Unclassified
                                                                        22. PRICE
EPA Form 2220-1 (9-73)
                                            116
                                                                     S GOVERNMfWT PRINTING OFFICE 1978—657-060/1489

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